Celery cultivar TBG 40

ABSTRACT

A celery cultivar, designated TBG 40, is disclosed. The invention relates to the seeds of celery cultivar TBG 40, to the plants of celery cultivar TBG 40 and to methods for producing a celery plant by crossing the cultivar TBG 40 with itself or another celery cultivar. The invention further relates to methods for producing a celery plant containing in its genetic material one or more transgenes and to the transgenic celery plants and plant parts produced by those methods. This invention also relates to celery cultivars or breeding cultivars and plant parts derived from celery cultivar TBG 40, to methods for producing other celery cultivars, lines or plant parts derived from celery cultivar TBG 40 and to the celery plants, varieties, and their parts derived from the use of those methods. The invention further relates to hybrid celery seeds, plants, and plant parts produced by crossing cultivar TBG 40 with another celery cultivar.

BACKGROUND OF THE INVENTION

The present invention relates to a new and distinctive celery (Apiumgraveolens var. dulce) variety, designated TBG 40. All publicationscited in this application are herein incorporated by reference.

There are numerous steps in the development of any novel, desirableplant germplasm. Plant breeding begins with the analysis, definition ofproblems and weaknesses of the current germplasm, the establishment ofprogram goals, and the definition of specific breeding objectives. Thenext step is selection of germplasm that possesses the traits to meetthe program goals. The goal is to combine in a single variety or hybridan improved combination of desirable traits from the parental germplasm.These important traits may include improved flavor, increased stalk sizeand weight, higher seed yield, improved color, resistance to diseasesand insects, tolerance to drought and heat, and better agronomicquality.

All cultivated forms of celery belong to the species Apium graveolensvar. dulce that is grown for its edible stalk. As a crop, celery isgrown commercially wherever environmental conditions permit theproduction of an economically viable yield. In the United States, theprincipal growing regions are California, Florida, Arizona and Michigan.Fresh celery is available in the United States year-round, although thegreatest supply is from November through January. For planting purposes,the celery season is typically divided into two seasons: summer andwinter, with Florida and the southern California areas harvesting fromNovember to July, and Michigan and northern California harvesting fromJuly to October. Celery is consumed as fresh, raw product and as acooked vegetable.

Celery is a cool-season biennial that grows best from 60° F. to 65° F.(16° C. to 18° C.), but will tolerate temperatures from 45° F. to 75° F.(7° C. to 24° C.). Freezing damages mature celery by splitting thepetioles or causing the skin to peel, making the stalks unmarketable.This can be a problem for crops planted in the winter regions; however,celery can tolerate minor freezes early in the production cycle.

The two main growing regions for celery in California are located alongthe Pacific Ocean: the central coast or summer production area(Monterey, San Benito, Santa Cruz and San Luis Obispo Counties) and thesouth coast or winter production area (Ventura and Santa BarbaraCounties). A minor region (winter) is located in the southern deserts(Riverside and Imperial Counties).

In the south coast, celery is transplanted from early August to Aprilfor harvest from November to mid-July; in the Santa Maria area, celeryis transplanted from January to August for harvest from April throughDecember. In the central coast, fields are transplanted from March toSeptember for harvest from late June to late December. In the southerndeserts, fields are transplanted in late August for harvest in January.

Commonly used celery varieties for coastal production include Tall Utah52-75, Conquistador and Sonora. Some shippers use their own proprietaryvarieties. Celery seed is very small and difficult to germinate. Allcommercial celery is planted as greenhouse-grown transplants. Celerygrown from transplants is more uniform than from seed and takes lesstime to grow the crop in the field. Transplanted celery is traditionallyplaced in double rows on 40-inch (100-cm) beds with plants spacedbetween 6.0 and 7.0 inches apart.

Celery requires a relatively long and cool growing season (Thephysiology of vegetable crops by Pressman, CAB Intl., New York, 1997).Earlier transplanting results in a longer growing season, increasedyields, and better prices. However, celery scheduled for spring harvestoften involves production in the coolest weather conditions of winter, aperiod during which vernalization can occur. If adequate vernalizationoccurs for the variety, bolting may be initiated. Bolting is thepremature rapid elongation of the main celery stem into a floral axis(i.e., during the first year for this normally biennial species).Bolting slows growth as the plant approaches marketable size and leavesa stalk with no commercial value. Different varieties have differentvernalization requirements, but in the presence of bolting, the lengthof the seed stem can be used as a means of measuring bolting tolerancethat exists in each different variety. The most susceptible varietiesreach their vernalization requirement earlier and have time to developthe longest seed stems, while the moderately tolerant varieties takelonger to reach their vernalization requirement and have less time todevelop a seed stem which would therefore be shorter. Under normalproduction conditions, the most tolerant varieties may not achieve theirvernalization requirement and therefore not produce a measurable seedstem.

The coldest months when celery is grown in the United States areDecember, January and February. If celery is going to reach itsvernalization requirements to cause bolting, it is generally youngercelery that is exposed to this cold weather window. This celerygenerally matures in the months of April and May which constitutes whatthe celery industry calls the bolting or seeder window. The bolting orseeder window is a period where seed stems are generally going to impactthe quality of the marketable celery and this is most consistentlyexperienced in celery grown in the Southern California region. Thepresence of seed stems in celery can be considered a major marketabledefect as set forth in the USDA grade standards. If the seed stem islonger than twice the diameter of the celery stalk or eight inches, thecelery no longer meets the standards of US Grade #1. If the seed stem islonger than three times the diameter of the celery stalk, the celery isno longer marketable as US Grade #2 (United States Standards for Gradesof Celery, United States Department of Agriculture, reprinted January1997).

Celery is an allogamous biennial crop. The celery genome consists of 11chromosomes. Its high degree of out-crossing is accomplished by insectsand wind pollination. Pollinators of celery flowers include a largenumber of wasp, bee and fly species. Celery is subject to inbreedingdepression, which appears to be dependent upon the genetic background assome lines are able to withstand selfing for three or four generations.

Celery flowers are protandrous, with pollen being released 3-6 daysbefore stigma receptivity. At the time of stigma receptivity the stamenswill have fallen and the two stigmata will have unfolded in an uprightposition. The degree of protandry varies, which makes it difficult toperform reliable hybridization, due to the possibility of accidentalselfing.

Celery flowers are very small, which significantly hinders easy removalof individual anthers. Furthermore, different developmental stages ofthe flowers in umbels make it difficult to avoid uncontrolledpollinations. The standard hybridization technique in celery consists ofselecting flower buds of the same size and eliminating the older andyounger flowers. Then, the umbellets are covered with glycine paper bagsfor a 5-10 day period, during which the stigmas become receptive. At thetime the flowers are receptive, available pollen or umbellets sheddingpollen from selected male parents are rubbed on to the stigmas of thefemale parent.

Celery plants require a period of vernalization while in the vegetativephase in order to induce seed stalk development. A period of 6-10 weeksat 5° C. to 8° C. when the plants are greater than 4 weeks old isusually adequate for most non-bolting tolerant varieties. Due to a widerange of responses to the cold treatment, it is often difficult tosynchronize crossing, since plants will flower at different times.However, pollen can be stored for 6-8 months at −10° C. in the presenceof silica gel or calcium chloride with a viability decline of only20-40%, thus providing flexibility to perform crosses over a longertime.

For selfing, the plant or selected umbels are caged in cloth bags. Theseare shaken several times during the day to promote pollen release.Houseflies (Musca domestica) can also be introduced weekly into the bagsto perform pollinations.

Celery in general is an important and valuable vegetable crop. Thus, acontinuing goal of celery plant breeders is to develop stable, highyielding celery cultivars that are resistant to diseases andagronomically sound to maximize the amount of yield produced on theland. To accomplish this goal, the celery breeder must select anddevelop celery plants that have the traits that result in superiorcultivars.

The foregoing examples of the related art and limitations relatedtherewith are intended to be illustrative and not exclusive. Otherlimitations of the related art will become apparent to those of skill inthe art upon a reading of the specification.

SUMMARY OF THE INVENTION

The following embodiments and aspects thereof are described inconjunction with systems, tools, and methods which are meant to beexemplary and illustrative, not limiting in scope. In variousembodiments, one or more of the above-described problems have beenreduced or eliminated, while other embodiments are directed to otherimprovements.

According to the invention, there is provided a novel celery cultivardesignated TBG 40. Also provided are celery plants having thephysiological and morphological characteristics of celery cultivar TBG40. This invention thus relates to the seeds of celery cultivar TBG 40,to the plants of celery cultivar TBG 40, and to methods for producing acelery plant by crossing celery TBG 40 with itself or another celeryplant, to methods for producing a celery plant containing in its geneticmaterial one or more transgenes and to the transgenic celery plantsproduced by that method. This invention also relates to methods forproducing other celery cultivars derived from celery cultivar TBG 40 andto the celery cultivar derived by the use of those methods. Thisinvention further relates to hybrid celery seeds and plants produced bycrossing celery cultivar TBG 40 with another celery line.

This invention further relates to the F₁ hybrid celery plants and plantparts grown from the hybrid seed produced by crossing celery cultivarTBG 40 to a second celery plant. Still further included in the inventionare the seeds of an F₁ hybrid plant produced with the celery cultivarTBG 40 as one parent, the second generation (F₂) hybrid celery plantgrown from the seed of the F₁ hybrid plant, and the seeds of the F₂hybrid plant. Thus, any such methods using the celery cultivar TBG 40are part of this invention: selfing, backcrosses, hybrid production,crosses to populations, and the like. All plants produced using celerycultivar TBG 40 as at least one parent are within the scope of thisinvention. Advantageously, the celery cultivar could be used in crosseswith other, different, celery plants to produce first generation (F₁)celery hybrid seeds and plants with superior characteristics.

The invention further provides methods for developing celery plantsderived from celery cultivar TBG 40 in a celery plant breeding programusing plant breeding techniques including recurrent selection,backcrossing, pedigree breeding, restriction fragment lengthpolymorphism enhanced selection, genetic marker enhanced selection andtransformation. Seeds, celery plants, and parts thereof, produced bysuch breeding methods are also part of the invention.

In another aspect, the present invention provides protoplasts andregenerable cells for use in tissue culture of celery cultivar TBG 40.The tissue culture will preferably be capable of regenerating plantshaving essentially all of the physiological and morphologicalcharacteristics of the foregoing celery plant, and of regeneratingplants having substantially the same genotype as the foregoing celeryplant. Preferably, the regenerable cells in such tissue cultures will becallus, protoplasts, meristematic cells, leaves, pollen, embryos, roots,root tips, anthers, pistils, flowers, seeds, petioles and suckers. Stillfurther, the present invention provides celery plants regenerated fromthe tissue cultures of the invention.

In another aspect, the present invention provides for single or multiplegene converted plants of TBG 40. The single or multiple transferredgene(s) may preferably be a dominant or recessive allele. Preferably,the single or multiple transferred gene(s) will confer such traits asmale sterility, herbicide resistance, insect resistance, modified fattyacid metabolism, modified carbohydrate metabolism, resistance forbacterial, fungal, or viral disease, male fertility, enhancednutritional quality and industrial usage or the transferred gene willhave no apparent value except for the purpose of being a marker forvariety identification. The single or multiple gene(s) may be anaturally occurring celery gene or a transgene introduced throughgenetic engineering techniques.

The invention also relates to methods for producing a celery plantcontaining in its genetic material one or more transgenes and to thetransgenic celery plant produced by those methods.

In another aspect, the present invention provides for methods ofintroducing one or more desired trait(s) into celery cultivar TBG 40 andplants or seeds obtained from such methods. The desired trait(s) may be,but not exclusively, a single gene, preferably a dominant but also arecessive allele. Preferably, the transferred gene or genes will confersuch traits as male sterility, herbicide resistance, insect resistance,disease resistance, resistance for bacterial, fungal, or viral disease,male fertility, water stress tolerance, enhanced nutritional quality,modified fatty acid metabolism, modified carbohydrate metabolism,enhanced plant quality, and industrial usage. The gene or genes may benaturally occurring rice gene(s). The method for introducing the desiredtrait(s) may be a backcrossing process making use of a series ofbackcrosses to the celery cultivar TBG 40 during which the desiredtrait(s) is maintained by selection. The desired trait may also beintroduced via transformation.

The invention also relates to methods for genetically modifying a celeryplant of celery cultivar TBG 40 and to the modified celery plantproduced by those methods. The genetic modification methods may include,but are not limited to mutation, genome editing, gene silencing, RNAinterference, backcross conversion, genetic transformation, single andmultiple gene conversion, and/or direct gene transfer. The inventionfurther relates to a genetically modified celery plant produced by theabove methods, wherein the genetically modified celery plant comprisesthe genetic modification and otherwise comprises essentially all of thephysiological and morphological characteristics of celery cultivar TBG40. In another aspect, the invention relates to a genetically modifiedcelery plant produced by the above methods, wherein the geneticallymodified celery plant comprises said mutation, genome editing, genesilencing, RNA interference, backcross conversion, genetictransformation, single and multiple gene conversion, and/or direct genetransfer and otherwise comprises essentially all of the physiologicaland morphological characteristics of celery cultivar TBG 40.

In addition to the exemplary aspects and embodiments described above,further aspects and embodiments will become apparent by reference by thestudy of the following descriptions.

DETAILED DESCRIPTION OF THE INVENTION

In the description and tables which follow, a number of terms are used.In order to provide a clear and consistent understanding of thespecification and claims, including the scope to be given such terms,the following definitions are provided:

Allele. An allele is any of one or more alternative form of a gene, allof which relate to one trait or characteristic. In a diploid cell ororganism, the two alleles of a given gene occupy corresponding loci on apair of homologous chromosomes.

Backcrossing. Backcrossing is a process in which a breeder repeatedlycrosses hybrid progeny back to one of the parents, for example, a firstgeneration hybrid F₁ with one of the parental genotype of the F₁ hybrid.

Bacterial blight. A bacterial disease of celery caused by Pseudomonassyringae pv. apii. The initial symptoms appear on the leaves as small,bright yellow, circular spots. As these enlarge with a yellow halo, theyturn to a rust color. As the spots increase in number they merge toeventually kill the leaf tissue. Bacterial blight is favored by cool,wet conditions and at least 10 hours of leaf wetness is required forinfection. The disease is spread by water splashes, farm machinery andfield workers especially when the foliage is wet.

Black streak. A physiological disorder in celery plants causing somepetioles, when cut, to show “black streaks” in the lower half orthroughout the entire length of the petiole, making the entire cropunmarketable. Symptoms can be triggered under field conditions by hightemperatures.

Blackheart. Blackheart is due to a lack of movement of sufficientcalcium that causes the plant to turn brown and begin to decay at thegrowing point of the plant. Celery in certain conditions, such as warmweather, grows very rapidly and is incapable of moving sufficientamounts of calcium to the growing point.

Bolting. The premature development of a flowering or seed stalk, andsubsequent seed, before a plant produces a food crop. Bolting istypically caused by late planting when temperatures are low enough tocause vernalization of the plants.

Bolting period. Also known as the bolting or seeder window, andgenerally occurs in celery that is transplanted from the middle ofDecember through January and matures in April to May. The intensity andactual weeks that bolting may be observed vary from year to year, but itis generally observed in this window.

Bolting tolerance. The amount of vernalization that is required fordifferent celery varieties to bolt is genetically controlled. Varietieswith increased tolerance to bolting require greater periods ofvernalization in order to initiate bolting. A comparison of boltingtolerance between varieties can be measured by the length of theflowering or seed stem under similar vernalization conditions.

Brown stem. A disease caused by the bacterium Pseudomonas cichorii thatcauses petiole necrosis. Brown Stem is characterized by a firm, browndiscoloration throughout the petiole.

Celeriac or Root celery (Apium graveolens L. var. rapaceum). A plantthat is related to celery but instead of having a thickened andsucculent leaf petiole as in celery, celeriac has an enlarged hypocotyland upper root that is the edible product.

Celery heart. The center most interior petioles and leaves of the celerystalk. They are not only the smallest petioles in the stalk, but theyoungest as well. Some varieties are considered heartless because theygo right from very large petioles to only a couple of very smallpetioles. The heart is comprised of the petioles that are closest to themeristem of the celery stalk.

Colletotrichum. One of the most common and important genera ofplant-pathogenic fungi. Causes post-harvest rots, and anthracnose spotsand blights of aerial plant parts. In celery it is also frequentlyaccompanied by curling of the foliage and black heart.

Consumable. Means material that is edible by humans.

Crackstem. The petiole can crack or split horizontally orlongitudinally. Numerous cracks in several locations along the petioleare often an indication that the variety has insufficient boronnutrition. A variety's ability to utilize boron is a physiologicalcharacteristic which is genetically controlled.

Essentially all of the physiological and morphological characteristics.A plant having essentially all of the physiological and morphologicalcharacteristics of a designated plant has all of the characteristics ofthe plant that are otherwise present when compared in the sameenvironment, other than an occasional variant trait that might ariseduring backcrossing, direct introduction of a transgene, or geneticmodification.

F_(#). The “F” symbol denotes the filial generation, and the # is thegeneration number, such as F₁, F₂, F₃, etc.

F₁ hybrid. The first generation progeny of the cross of two nonisogenicplants.

Feather leaf. Feather leaf is a yellowing of the lower leaflets andgenerally occurs in the outer petioles but can also be found on innerpetioles of the stalk. These yellowing leaves which would normallyremain in the harvested stalk are considered unacceptable. Thesepetioles then have to be stripped off in order to meet USDA standardswhich effectively decreases the stalk size and yield.

Flare. The lower, generally wider portion of the petiole which isusually a paler green or white. Some also refer to this as the spoon,scoop, or shovel.

Fusarium yellows. A fungal soilborne disease caused by Fusariumoxysporum f sp. apii Race 2. Infected plants turn yellow and arestunted. Some of the large roots may have a dark brown and awater-soaked appearance. The water-conducting tissue (xylem) in thestem, crown, and root show a characteristic orange-brown discoloration.In the later stages of infection, plants remain severely stunted andyellowed and may collapse. The disease appears most severe during warmseasons, and in heavy, wet soils.

Gene. As used herein, “gene” refers to a segment of nucleic acid. A genecan be introduced into a genome of a species, whether from a differentspecies or from the same species, using transformation or variousbreeding techniques.

Gene silencing. The interruption or suppression of the expression of agene at the level of transcription or translation.

Genetically modified. Describes an organism that has received geneticmaterial from another organism, or had its genetic material modified,resulting in a change in one or more of its phenotypic characteristics.Methods used to modify, introduce or delete the genetic material mayinclude mutation breeding, genome editing, RNA interference, backcrossconversion, genetic transformation, single and multiple gene conversion,and/or direct gene transfer.

Genome editing. A type of genetic engineering in which DNA is inserted,replaced, modified or removed from a genome using artificiallyengineered nucleases or other targeted changes using homologousrecombination. Examples include but are not limited to use of zincfinger nucleases (ZFNs), TAL effector nucleases (TALENs), endonucleases,meganucleases, CRISPR/Cas9, and other CRISPR related technologies. (Maet. al., Molecular Plant, 9:961-974 (2016); Belhaj et. al., CurrentOpinion in Biotechnology, 32:76-84 (2015)).

Gross yield (Pounds/Acre). The total yield in pounds/acre of trimmedcelery plants (stalks).

Leaf celery (Apium graveolens L. var. secalinum). A plant that has beendeveloped primarily for leaf and seed production. Often grown inMediterranean climates, leaf celery more closely resembles celery's wildancestors. The stems are small and fragile and vary from solid to hollowand the leaves are fairly small and are generally bitter. This type isoften used for its medicinal properties and spice.

Leaf margin chlorosis. A magnesium deficiency producing an interveinalchlorosis which starts at the margin of leaves.

Maturity date. Maturity in celery can be dictated by two conditions. Thefirst, or true maturity, is the point in time when the celery reachesmaximum size distribution, but before defects such as pith, yellowing,Feather Leaf or Brown Stem appear. The second, or market maturity is anartificial maturity dictated by market conditions, i.e, the marketrequirement may be for 3 dozen sizes so the field is harvested atslightly below maximum yield potential because the smaller sizes arewhat the customers prefer at that moment.

Muck. Muck is a soil made up primarily of humus drained from swampland.It is used for growing specialty crops, such as onions, carrots, celery,and potatoes.

MUN. MUN refers to the MUNSELL Color Chart which publishes an officialcolor chart for plant tissues according to a defined numbering system.The chart may be purchased from the Macbeth Division of KollmorgenInstruments Corporation, 617 Little Britain Road, New Windsor, N.Y.12553-6148.

Petiole. A petiole is the stem or limb of a leaf, the primary portion ofthe celery consumed.

Petiole depth. The average measurement in millimeters of the depth ofthe celery petiole at its narrowest point. The petiole depth measurementis taken from the outside of the petiole (which is the part of thepetiole that faces the outside of the stalk) and is measured to theinside of the petiole or cup or the inner most point of the petiole thatfaces the center of the stalk or heart.

Petiole width. The average measurement of the width of the celerypetiole in millimeters at its widest point. The measurement is takenfrom the side or edge of petiole to the opposite side or edge of thepetiole. The measurement is taken 90 degrees from petiole depth.

Phthalides. One of the chemical compounds that are responsible for thecharacteristic flavor and aroma of celery.

Pith. Pith is a sponginess/hollowness/white discoloration that occurs inthe petioles of celery varieties naturally as they become over-mature.In some varieties it occurs at an earlier stage causing harvest to occurprior to ideal maturity. Pith generally occurs in the outer, olderpetioles first. If it occurs, these petioles are stripped off to makegrade, which effectively decreases the stalk size and overall yieldpotential.

Plant height. The height of the plant from the bottom of the base orbutt of the celery plant to the top of the tallest leaf.

Polyphenol oxidase (PPO). An enzyme that catalyzes the conversion ofphenolic compounds to quinones and assists their products'polymerization. The catalysis of PPO, in the presence of oxygen, leadsto the formation of undesirable brown pigments and off-flavoredproducts.

Quantitative Trait Loci. Quantitative Trait Loci (QTL) refers to geneticloci that control to some degree, numerically representable traits thatare usually continuously distributed.

Regeneration. Regeneration refers to the development of a plant fromtissue culture.

Ribbing. The texture of the exterior surface of the celery petiole canvary from smooth to ribby depending on the variety. Ribbing is thepresence of numerous ridges that run vertically along the petioles ofthe celery plant.

Seed stem. A seed stem is the result of the elongation of the main stemof the celery, which is usually very compressed to almost non-existent,to form the flowering axis. The seed stem or flowering axis can reachseveral feet in height during full flower. The length of the seed stemis measured as the distance from the top of the basal plate (the base ofthe seed stem) to its terminus (the terminal growing point). Septoriaapiicola. A fungus that is the cause of late blight in celery. Symptomsinclude chlorotic spots that turn brown and necrotic.

Single gene converted. Single gene converted or conversion plant refersto plants which are developed by backcrossing, or via geneticengineering, wherein essentially all of the desired morphological andphysiological characteristics of a line are recovered in addition to thesingle gene transferred into the line via the backcrossing technique orvia genetic engineering.

Stalk. A stalk is a single celery plant that is trimmed with the top orfoliage and the roots removed.

Standard stem celery. A more traditional stem celery with moderate jointlength, to be utilized and marketed as a whole stalk with 12 to 14 inchcut or for hearts in retail environment.

Stringiness. Stringiness is a physiological characteristic that isgenerally associated with strings that get stuck between the consumer'steeth. There are generally two sources of strings in celery. One is thevascular bundle which can be fairly elastic and behave as a string. Thesecond is a strip of particularly strong epidermis cells calledschlerenchyma which are located on the surface of the ridges of thecelery varieties that have ribs.

Suckers. Suckers are auxiliary shoots that form at the base of the stalkor within the auxiliary buds between each petiole. If these shoots formbetween the petioles of the stalk, several petioles have to be strippedoff causing the celery to become smaller and the functional yields to bedecreased.

Tall stem celery. A stem celery with especially long petioles withprimary purpose of being utilized for production of sticks or limbs.

Transgene. A nucleic acid of interest that can be introduced into thegenome of a plant by genetic engineering techniques (e.g.,transformation) or breeding.

Breeding History of Celery Cultivar TBG 40

The development of celery cultivar TBG 40 began in 2001 when severaldwarf mutants appeared in a commercial crop of celery variety Hill'sSpecial (U.S. PVP #9500019) that were produced from an aged seed lotwhich was being transitioned out of use due to poor seed quality. Thedwarf mutants appeared at a rate of approximately 1 in 80,000 plants.Five dwarf mutants were selected and selfed. These mutants reproduced aspure true breeding dwarfs and had very similar appearances. Theinheritance of the dwarf trait appears to be a single recessive gene.When the dwarf was crossed with a normal celery, the F₁ progeny was 100%normal and the F₂ progeny produced approximately 25% dwarf celery and75% normal celery. These mutants were never found in healthy seed lotsof Hill's Special. One of the five original dwarf selfs was placed intoseed production and the result was the uniform and consistent, truebreeding celery variety designated TBG 40.

Celery cultivar TBG 40 has the following morphologic and othercharacteristics (based primarily on data collected in California):

TABLE 1 VARIETY DESCRIPTION INFORMATION Maturity: 100 days Plant height:41.5 cm Whole plant weight: 0.69 kg Trimmed plant weight: 0.62 kg Numberof Outer Petioles: 13.4 Number of Inner Petioles: 7.4 Stalk shape:Cylindrical Stalk conformation: Compact Heart formation: The stalk has afull heart, but the stalk is essentially a heart in its own right Lengthof outer petioles 12.8 cm (from butt to first joint): Width of outerpetioles (at midpoint): 29.5 mm Thickness of outer petioles 8.6 mm (atmidpoint): Petiole length class: Short Petiole cross section shape:Slight cup Petiole color: MUN 5 gy 6/6 Anthocyanin: None Stringiness:Normal Ribbing: Smooth Leaf blade color: MUN 5 gy 3/4 Bolting tolerance:Moderate Stress tolerance: Adaxial Crackstem (Boron Deficiency): NormalAbaxial Crackstem (Boron Deficiency): Normal Leaf Margin ChlorosisTolerant (Magnesium Deficiency): Blackheart (Calcium Deficiency):Tolerant Pithiness(Nutritional Deficiency): Normal Feather Leaf:Tolerant Sucker Development: Moderately tolerant Disease resistance:Brown Stem (Pseudomonas cichorii): Moderately tolerant Bacterial Blight(Pseudomonas Unknown syringae pv. apii): Late Blight (Septoria apii):Susceptible Fusarium oxysporum f. sp. apii race 2: Susceptible

This invention is also directed to methods for producing a celery plantby crossing a first parent celery plant with a second parent celeryplant, wherein the first parent celery plant or second parent celeryplant is celery cultivar TBG 40. Further, both the first parent celeryplant and second parent celery plant may be from celery cultivar TBG 40.Still further, this invention also is directed to methods for producinga cultivar TBG 40-derived celery plant by crossing cultivar TBG 40 witha second celery plant and growing the progeny seed, and repeating thecrossing and growing steps with the cultivar TBG 40-derived plant from 0to 7 times. Thus, any such methods using the cultivar TBG 40 are part ofthis invention: selfing, backcrosses, hybrid production, crosses topopulations, and the like. All plants produced using cultivar TBG 40 asa parent are within the scope of this invention, including plantsderived from cultivar TBG 40. Advantageously, cultivar TBG 40 can beused in crosses with other, different, cultivars to produce firstgeneration (F₁) celery seeds and plants with superior characteristics.

Additional methods include, but are not limited to, expression vectorsintroduced into plant tissues using a direct gene transfer method suchas microprojectile-mediated delivery, DNA injection, electroporation andthe like. More preferably, expression vectors are introduced into planttissues by using either microprojectile-mediated delivery with abiolistic device or by using Agrobacterium-mediated transformation.Transformant plants obtained with the protoplasm of the invention areintended to be within the scope of this invention.

As used herein, the term “plant” includes plant cells, plantprotoplasts, plant cell tissue cultures from which celery plants can beregenerated, plant calli, plant clumps and plant cells that are intactin plants or parts of plants, such as leaves, pollen, embryos,meristematic cells, hypocotyls, roots, root tips, anthers, pistils,flowers, seeds, stems, and the like.

Further Embodiments of the Invention

Celery in general is an important and valuable vegetable crop. Thus, acontinuing goal of celery plant breeders is to develop stable, highyielding celery cultivars that are agronomically sound. To accomplishthis goal, the celery breeder must select and develop celery plants withtraits that result in superior cultivars.

Plant breeding techniques known in the art and used in a celery plantbreeding program include, but are not limited to, pedigree breeding,recurrent selection, mass selection, single or multiple-seed descent,bulk selection, backcrossing, open pollination breeding, restrictionfragment length polymorphism enhanced selection, genetic marker enhancedselection, making double haploids, and transformation. Oftencombinations of these techniques are used. The development of celeryvarieties in a plant breeding program requires, in general, thedevelopment and evaluation of homozygous varieties. There are manyanalytical methods available to evaluate a new variety. The oldest andmost traditional method of analysis is the observation of phenotypictraits, but genotypic analysis may also be used.

Using Celery Cultivar TBG 40 to Develop Other Celery Varieties

This invention also is directed to methods for producing a celery plantby crossing a first parent celery plant with a second parent celeryplant wherein the first or second parent celery plant is a celery plantof cultivar TBG 40. Further, both first and second parent celery plantscan come from celery cultivar TBG 40. Also provided are methods forproducing a celery plant having substantially all of the morphologicaland physiological characteristics of cultivar TBG 40, by crossing afirst parent celery plant with a second parent celery plant wherein thefirst and/or the second parent celery plant is a plant havingsubstantially all of the morphological and physiological characteristicsof cultivar TBG 40 as determined at the 5% significance level when grownin the same environmental conditions. The other parent may be any celeryplant, such as a celery plant that is part of a synthetic or naturalpopulation. Thus, any such methods using celery cultivar TBG 40 are partof this invention: selfing, backcrosses, hybrid production, crosses topopulations, and the like. All plants produced using celery cultivar TBG40 as at least one parent are within the scope of this invention,including those developed from cultivars derived from celery cultivarTBG 40.

The cultivar of the invention can also be used for transformation whereexogenous genes are introduced and expressed by the cultivar of theinvention. Genetic variants created either through traditional breedingmethods using celery cultivar TBG 40 or through transformation ofcultivar TBG 40 by any of a number of protocols known to those of skillin the art are intended to be within the scope of this invention.

The following describes breeding methods that may be used with celerycultivar TBG 40 in the development of further celery plants. One suchembodiment is a method for developing a progeny celery plant in a celeryplant breeding program comprising: obtaining the celery plant, or a partthereof, of cultivar TBG 40, utilizing said plant or plant part as asource of breeding material, and selecting a celery cultivar TBG 40progeny plant with molecular markers in common with cultivar TBG 40and/or with morphological and/or physiological characteristics of celerycultivar TBG 40. Breeding steps that may be used in the celery plantbreeding program include, but are not limited to, pedigree breeding,backcrossing, mutation breeding, and recurrent selection. In conjunctionwith these steps, techniques such as RFLP-enhanced selection, geneticmarker enhanced selection (for example SSR markers) and the making ofdouble haploids may be utilized.

Another method involves producing a population of celery cultivar TBG 40progeny celery plants, comprising crossing cultivar TBG 40 with anothercelery plant, thereby producing a population of celery plants, which, onaverage, derive 50% of their alleles from celery cultivar TBG 40. Aplant of this population may be selected and repeatedly selfed or sibbedwith a celery cultivar resulting from these successive filialgenerations. One embodiment of this invention is the celery cultivarproduced by this method and that has obtained at least 50% of itsalleles from celery cultivar TBG 40.

Still yet another aspect of the invention is a method of producing acelery plant derived from the celery cultivar TBG 40, the methodcomprising the steps of: (a) preparing a progeny plant derived fromcelery cultivar TBG 40 by crossing a plant of the celery cultivar TBG 40with a second celery plant; and (b) crossing the progeny plant withitself or a second plant to produce a seed of a progeny plant of asubsequent generation which is derived from a plant of the celerycultivar TBG 40. In further embodiments of the invention, the method mayadditionally comprise: (c) growing a progeny plant of a subsequentgeneration from said seed of a progeny plant of a subsequent generationand crossing the progeny plant of a subsequent generation with itself ora second plant; and repeating the steps for an additional 2-10generations to produce a celery plant derived from the celery cultivarTBG 40. The plant derived from celery cultivar TBG 40 may be an inbredline, and the aforementioned repeated crossing steps may be defined ascomprising sufficient inbreeding to produce the inbred line. In themethod, it may be desirable to select particular plants resulting fromstep (c) for continued crossing according to steps (b) and (c). Byselecting plants having one or more desirable traits, a plant derivedfrom celery cultivar TBG 40 is obtained which possesses some of thedesirable traits of the line as well as potentially other selectedtraits. Also provided by the invention is a plant produced by this andthe other methods of the invention.

In another embodiment of the invention, the method of producing a celeryplant derived from the celery cultivar TBG 40 further comprises: (a)crossing the celery cultivar TBG 40-derived celery plant with itself oranother celery plant to yield additional celery cultivar TBG 40-derivedprogeny celery seed; (b) growing the progeny celery seed of step (a)under plant growth conditions to yield additional celery cultivar TBG40-derived celery plants; and (c) repeating the crossing and growingsteps of (a) and (b) to generate further celery cultivar TBG 40-derivedcelery plants. In specific embodiments, steps (a) and (b) may berepeated at least 1, 2, 3, 4, or 5 or more times as desired. Theinvention still further provides a celery plant produced by this and theforegoing methods.

Progeny of celery cultivar TBG 40 may also be characterized throughtheir filial relationship with celery cultivar TBG 40, as for example,being within a certain number of breeding crosses of celery cultivar TBG40. A breeding cross is a cross made to introduce new genetics into theprogeny, and is distinguished from a cross, such as a self or a sibcross, made to select among existing genetic alleles. The lower thenumber of breeding crosses in the pedigree, the closer the relationshipbetween celery cultivar TBG 40 and its progeny. For example, progenyproduced by the methods described herein may be within 1, 2, 3, 4 or 5breeding crosses of celery cultivar TBG 40.

One of ordinary skill in the art of plant breeding would know how toevaluate the traits of two plant varieties to determine if there is nosignificant difference between the two traits expressed by thosevarieties. For example, see, Fehr and Walt, Principles of CultivarDevelopment, pp. 261-286 (1987). Thus the invention includes celerycultivar TBG 40 progeny celery plants comprising a combination of atleast two cultivar TBG 40 traits selected from the group consisting ofthose listed in Table 1 or the cultivar TBG 40 combination of traitslisted in the Tables, so that said progeny celery plant is notsignificantly different for said traits than celery cultivar TBG 40 asdetermined at the 5% significance level when grown in the sameenvironmental conditions. Using techniques described herein, molecularmarkers may be used to identify said progeny plant as a celery cultivarTBG 40 progeny plant. Mean trait values may be used to determine whethertrait differences are significant, and preferably the traits aremeasured on plants grown under the same environmental conditions. Oncesuch a variety is developed its value is substantial since it isimportant to advance the germplasm base as a whole in order to maintainor improve traits such as yield, disease resistance, pest resistance,and plant performance in extreme environmental conditions.

The goal of celery plant breeding is to develop new, unique, andsuperior celery cultivars. The breeder initially selects and crosses twoor more parental lines, followed by repeated selfing and selection,producing many new genetic combinations. The breeder can theoreticallygenerate billions of different genetic combinations via crossing,selfing, and mutations. The breeder has no direct control at thecellular level and the cultivars that are developed are unpredictable.This unpredictability is because the breeder's selection occurs inunique environments, with no control at the DNA level (usingconventional breeding procedures), and with millions of differentpossible genetic combinations being generated. A breeder of ordinaryskill in the art cannot predict the final resulting lines he develops,except possibly in a very gross and general fashion. The same breedercannot produce the same line twice by using the exact same originalparents and the same selection techniques. Therefore, two breeders willnever develop the same line, or even very similar lines, having the samecelery traits.

Pedigree breeding is used commonly for the improvement ofself-pollinating crops or inbred lines of cross-pollinating crops.Pedigree breeding starts with the crossing of two genotypes, such ascelery cultivar TBG 40 or a celery variety having all of themorphological and physiological characteristics of TBG 40, and anothercelery variety having one or more desirable characteristics that islacking or which complements celery cultivar TBG 40. If the two originalparents do not provide all the desired characteristics, other sourcescan be included in the breeding population. In the pedigree method,superior plants are selfed and selected in successive filialgenerations. In the succeeding filial generations, the heterozygouscondition gives way to the homozygous allele condition as a result ofinbreeding. Typically in the pedigree method of breeding, five or moresuccessive filial generations of selfing and selection is practiced: F₁to F₂; F₂ to F₃; F₃ to F₄; F₄ to F₅; etc. In some examples, 1, 2, 3, 4,5, 6, 7, 8, 9, 10, or more generations of selfing and selection arepracticed. After a sufficient amount of inbreeding, successive filialgenerations will serve to increase seed of the developed variety.Preferably, the developed variety comprises homozygous alleles at about95% or more of its loci.

In addition to being used to create backcross conversion populations,backcrossing can also be used in combination with pedigree breeding. Asdiscussed previously, backcrossing can be used to transfer one or morespecifically desirable traits from one variety (the donor parent) to adeveloped variety (the recurrent parent), which has good overallagronomic characteristics yet may lack one or more other desirabletraits. However, the same procedure can be used to move the progenytoward the genotype of the recurrent parent but at the same time retainmany components of the non-recurrent parent by stopping the backcrossingat an early stage and proceeding with selfing and selection. Forexample, a celery variety may be crossed with another variety to producea first generation progeny plant. The first generation progeny plant maythen be backcrossed to one of its parent varieties to create a F₁BC₁.Progeny are selfed and selected so that the newly developed variety hasmany of the attributes of the recurrent parent and yet several of thedesired attributes of the donor parent. This approach leverages thevalue and strengths of both parents for use in new celery varieties.

Therefore, in some examples a method of making a backcross conversion ofcelery cultivar TBG 40, comprising the steps of crossing a plant ofcelery cultivar TBG 40 or a celery variety having all of themorphological and physiological characteristics of TBG 40 with a donorplant possessing a desired trait to introduce the desired trait,selecting an F₁ progeny plant containing the desired trait, andbackcrossing the selected F₁ progeny plant to a plant of celery cultivarTBG 40 are provided. This method may further comprise the step ofobtaining a molecular marker profile of celery cultivar TBG 40 and usingthe molecular marker profile to select for a progeny plant with thedesired trait and the molecular marker profile of TBG 40. The molecularmarker profile can comprise information from one or more markers. In oneexample the desired trait is a mutant gene or transgene present in thedonor parent. In another example, the desired trait is a native trait inthe donor parent.

Mass and recurrent selections can be used to improve populations ofeither self- or cross-pollinating crops. A genetically variablepopulation of heterozygous individuals is either identified or createdby intercrossing several different parents. The best plants are selectedbased on individual superiority, outstanding progeny, or excellentcombining ability. The selected plants are intercrossed to produce a newpopulation in which further cycles of selection are continued.

The single-seed descent procedure in the strict sense refers to plantinga segregating population, harvesting a sample of one seed per plant, andusing the one-seed sample to plant the next generation. When thepopulation has been advanced from the F₂ to the desired level ofinbreeding, the plants from which lines are derived will each trace todifferent F₂ individuals. The number of plants in a population declineseach generation due to failure of some seeds to germinate or some plantsto produce at least one seed. As a result, not all of the F₂ plantsoriginally sampled in the population, will be represented by a progenywhen generation advance is completed.

Mutation breeding is another method of introducing new traits intocelery varieties. Mutations that occur spontaneously or are artificiallyinduced can be useful sources of variability for a plant breeder. Thegoal of artificial mutagenesis is to increase the rate of mutation for adesired characteristic. Mutation rates can be increased by manydifferent means including temperature, long-term seed storage, tissueculture conditions, radiation (such as X-rays, Gamma rays, neutrons,Beta radiation, or ultraviolet radiation), chemical mutagens (such asbase analogs like 5-bromo-uracil), antibiotics, alkylating agents (suchas sulfur mustards, nitrogen mustards, epoxides, ethyleneamines,sulfates, sulfonates, sulfones, or lactones), azide, hydroxylamine,nitrous acid or acridines. Once a desired trait is observed throughmutagenesis the trait may then be incorporated into existing germplasmby traditional breeding techniques. Details of mutation breeding can befound in “Principles of Cultivar Development” by Fehr, MacmillanPublishing Company, 1993. In addition, mutations created in other celeryplants may be used to produce a backcross conversion of celery cultivarTBG 40 that comprises such mutation.

Selection of celery plants for breeding is not necessarily dependent onthe phenotype of a plant and instead can be based on geneticinvestigations. For example, one may utilize a suitable genetic markerwhich is closely associated with a trait of interest. One of thesemarkers may therefore be used to identify the presence or absence of atrait in the offspring of a particular cross, and hence may be used inselection of progeny for continued breeding. This technique may commonlybe referred to as marker assisted selection. Any other type of geneticmarker or other assay which is able to identify the relative presence orabsence of a trait of interest in a plant may also be useful forbreeding purposes. Procedures for marker assisted selection applicableto the breeding of celery are well known in the art. Such methods willbe of particular utility in the case of recessive traits and variablephenotypes, or where conventional assays may be more expensive, timeconsuming or otherwise disadvantageous. Types of genetic markers whichcould be used in accordance with the invention include, but are notnecessarily limited to, Isozyme Electrophoresis, Restriction FragmentLength Polymorphisms (RFLPs), Simple Sequence Length Polymorphisms(SSLPs) (Williams et al., Nucleic Acids Res., 18:6531-6535, 1990),Randomly Amplified Polymorphic DNAs (RAPDs), DNA AmplificationFingerprinting (DAF), Sequence Characterized Amplified Regions (SCARs),Arbitrary Primed Polymerase Chain Reaction (AP-PCR), Amplified FragmentLength Polymorphisms (AFLPs) (EP 534 858, specifically incorporatedherein by reference in its entirety), Simple Sequence Repeats (SSRs),and Single Nucleotide Polymorphisms (SNPs) (Wang et al., Science,280:1077-1082, 1998).

The production of double haploids can also be used for the developmentof homozygous varieties in a breeding program. Double haploids areproduced by the doubling of a set of chromosomes from a heterozygousplant to produce a completely homozygous individual. For example, see,Wan, et al., “Efficient Production of Doubled Haploid Plants ThroughColchicine Treatment of Anther-Derived Maize Callus,” Theoretical andApplied Genetics, 77:889-892 (1989) and U.S. Pat. No. 7,135,615. Thiscan be advantageous because the process omits the generations of selfingneeded to obtain a homozygous plant from a heterozygous source.

Descriptions of other breeding methods that are commonly used fordifferent traits and crops can be found in one of several referencebooks (e.g., Allard, “Principles of plant breeding,” John Wiley & Sons,NY, University of California, Davis, Calif., 50-98, 1960; Simmonds,“Principles of crop improvement,” Longman, Inc., NY, 369-399, 1979;Sneep and Hendriksen, “Plant breeding perspectives,” Wageningen (ed),Center for Agricultural Publishing and Documentation, 1979; Fehr, In:Soybeans: Improvement, Production and Uses,” 2d Ed., Manograph 16:249,1987; Fehr, “Principles of cultivar development,” Theory and Technique(Vol 1) and Crop Species Soybean (Vol 2), Iowa State Univ., MacmillianPub. Co., NY, 360-376, 1987; Poehlman and Sleper, “Breeding Field Crops”Iowa State University Press, Ames, 1995; Sprague and Dudley, eds., Cornand Improvement, 5th ed., 2006).

Genotypic Profile of TBG 40 and Progeny

In addition to phenotypic observations, a plant can also be identifiedby its genotype. The genotype of a plant can be characterized through agenetic marker profile which can identify plants of the same variety ora related variety, or which can be used to determine or validate apedigree. Genetic marker profiles can be obtained by techniques such asrestriction fragment length polymorphisms (RFLPs), randomly amplifiedpolymorphic DNAs (RAPDs), arbitrarily primed polymerase chain reaction(AP-PCR), DNA amplification fingerprinting (DAF), sequence characterizedamplified regions (SCARs), amplified fragment length polymorphisms(AFLPs), simple sequence repeats (SSRs) also referred to asmicrosatellites, single nucleotide polymorphisms (SNPs), or genome-wideevaluations such as genotyping-by-sequencing (GBS). For example, seeCregan et al. (1999) “An Integrated Genetic Linkage Map of the SoybeanGenome” Crop Science 39:1464-1490, and Berry et al. (2003) “AssessingProbability of Ancestry Using Simple Sequence Repeat Profiles:Applications to Maize Inbred Lines and Soybean Varieties” Genetics165:331-342, each of which are incorporated by reference herein in theirentirety. Favorable genotypes and or marker profiles, optionallyassociated with a trait of interest, may be identified by one or moremethodologies.

In some examples one or more markers are used, including but not limitedto AFLPs, RFLPs, ASH, SSRs, SNPs, indels, padlock probes, molecularinversion probes, microarrays, sequencing, and the like. In somemethods, a target nucleic acid is amplified prior to hybridization witha probe. In other cases, the target nucleic acid is not amplified priorto hybridization, such as methods using molecular inversion probes (see,for example Hardenbol et al. (2003) Nat Biotech 21:673-678). In someexamples, the genotype related to a specific trait is monitored, whilein other examples, a genome-wide evaluation including but not limited toone or more of marker panels, library screens, association studies,microarrays, gene chips, expression studies, or sequencing such aswhole-genome resequencing and genotyping-by-sequencing (GBS) may beused. In some examples, no target-specific probe is needed, for exampleby using sequencing technologies, including but not limited tonext-generation sequencing methods (see, for example, Metzker (2010) NatRev Genet 11:31-46; and, Egan et al. (2012) Am J Bot 99:175-185) such assequencing by synthesis (e.g., Roche 454 pyrosequencing, Illumina GenomeAnalyzer, and Ion Torrent PGM or Proton systems), sequencing by ligation(e.g., SOLiD from Applied Biosystems, and Polnator system from AzcoBiotech), and single molecule sequencing (SMS or third-generationsequencing) which eliminate template amplification (e.g., Helicossystem, and PacBio RS system from Pacific BioSciences). Furthertechnologies include optical sequencing systems (e.g., Starlight fromLife Technologies), and nanopore sequencing (e.g., GridION from OxfordNanopore Technologies). Each of these may be coupled with one or moreenrichment strategies for organellar or nuclear genomes in order toreduce the complexity of the genome under investigation via PCR,hybridization, restriction enzyme (see, e.g., Elshire et al. (2011) PLoSONE 6:e19379), and expression methods. In some examples, no referencegenome sequence is needed in order to complete the analysis.

With any of the genotyping techniques mentioned herein, polymorphismsmay be detected when the genotype and/or sequence of the plant ofinterest is compared to the genotype and/or sequence of one or morereference plants. The polymorphism revealed by these techniques may beused to establish links between genotype and phenotype. Thepolymorphisms may thus be used to predict or identify certain phenotypiccharacteristics, individuals, or even species. The polymorphisms aregenerally called markers. It is common practice for the skilled artisanto apply molecular DNA techniques for generating polymorphisms andcreating markers. The polymorphisms of this invention may be provided ina variety of mediums to facilitate use, e.g. a database or computerreadable medium, which may also contain descriptive annotations in aform that allows a skilled artisan to examine or query the polymorphismsand obtain useful information.

The invention further provides a method of determining the genotype of aplant of celery cultivar TBG 40, or a first generation progeny thereof,comprising detecting in the genome of the plant at least a firstpolymorphism. The method may, in certain embodiments, comprise detectinga plurality of polymorphisms in the genome of the plant. The method mayfurther comprise storing the results of the step of detecting theplurality of polymorphisms on a computer readable medium. The pluralityof polymorphisms are indicative of and/or give rise to the expression ofthe morphological and physiological characteristics of celery cultivarTBG 40. The invention further provides a computer readable mediumproduced by such a method.

In some examples, a plant, a plant part, or a seed of celery cultivarTBG 40 may be characterized by producing a molecular profile. Amolecular profile may include, but is not limited to, one or moregenotypic and/or phenotypic profile(s). A genotypic profile may include,but is not limited to, a marker profile, such as a genetic map, alinkage map, a trait maker profile, a SNP profile, an SSR profile, agenome-wide marker profile, a haplotype, and the like. A molecularprofile may also be a nucleic acid sequence profile, and/or a physicalmap. A phenotypic profile may include, but is not limited to, a proteinexpression profile, a metabolic profile, an mRNA expression profile, andthe like.

SSR technology is currently the most efficient and practical markertechnology; more marker loci can be routinely used and more alleles permarker locus can be found using SSRs in comparison to RFLPs. Forexample, Diwan and Cregan described a highly polymorphic microsatellitelocus in soybean with as many as 26 alleles. Diwan, N. and Cregan, P.B., Theor. Appl. Genet., 95:22-225 (1997). SNPs may also be used toidentify the unique genetic composition of the invention and progenyvarieties retaining that unique genetic composition. Various molecularmarker techniques may be used in combination to enhance overallresolution.

Molecular markers, which include markers identified through the use oftechniques such as Isozyme Electrophoresis, RFLPs, RAPDs, AP-PCR, DAF,SCARs, AFLPs, SSRs, and SNPs, may be used in plant breeding. One use ofmolecular markers is Quantitative Trait Loci (QTL) mapping. QTL mappingis the use of markers which are known to be closely linked to allelesthat have measurable effects on a quantitative trait. Selection in thebreeding process is based upon the accumulation of markers linked to thepositive effecting alleles and/or the elimination of the markers linkedto the negative effecting alleles from the plant's genome.

Molecular markers can also be used during the breeding process for theselection of qualitative traits. For example, markers closely linked toalleles or markers containing sequences within the actual alleles ofinterest can be used to select plants that contain the alleles ofinterest during a backcrossing breeding program. The markers can also beused to select toward the genome of the recurrent parent and against themarkers of the donor parent. This procedure attempts to minimize theamount of genome from the donor parent that remains in the selectedplants. It can also be used to reduce the number of crosses back to therecurrent parent needed in a backcrossing program. The use of molecularmarkers in the selection process is often called genetic marker enhancedselection or marker-assisted selection. Molecular markers may also beused to identify and exclude certain sources of germplasm as parentalvarieties or ancestors of a plant by providing a means of trackinggenetic profiles through crosses.

Particular markers used for these purposes are not limited to the set ofmarkers disclosed herein, but may include any type of marker and markerprofile which provides a means of distinguishing varieties. In additionto being used for identification of celery cultivar TBG 40, a hybridproduced through the use of TBG 40, and the identification orverification of pedigree for progeny plants produced through the use ofTBG 40, a genetic marker profile is also useful in developing a geneconversion of TBG 40.

Means of performing genetic marker profiles using SNP and SSRpolymorphisms are well known in the art. SNPs are genetic markers basedon a polymorphism in a single nucleotide. A marker system based on SNPscan be highly informative in linkage analysis relative to other markersystems in that multiple alleles may be present.

The SSR profile of celery cultivar TBG 40 can be used to identify plantscomprising celery cultivar TBG 40 as a parent, since such plants willcomprise the same homozygous alleles as celery cultivar TBG 40. Becausethe celery variety is essentially homozygous at all relevant loci, mostloci should have only one type of allele present. In contrast, a geneticmarker profile of an F₁ progeny should be the sum of those parents,e.g., if one parent was homozygous for allele x at a particular locus,and the other parent homozygous for allele y at that locus, then the F₁progeny will be xy (heterozygous) at that locus. Subsequent generationsof progeny produced by selection and breeding are expected to be ofgenotype x (homozygous), y (homozygous), or xy (heterozygous) for thatlocus position. When the F₁ plant is selfed or sibbed for successivefilial generations, the locus should be either x or y for that position.

In addition, plants and plant parts substantially benefiting from theuse of celery cultivar TBG 40 in their development, such as celerycultivar TBG 40 comprising a gene conversion, backcross conversion,transgene, or genetic sterility factor, may be identified by having amolecular marker profile with a high percent identity to celery cultivarTBG 40. Such a percent identity might be 95%, 96%, 97%, 98%, 99%, 99.5%,or 99.9% identical to celery cultivar TBG 40.

The SSR profile of celery cultivar TBG 40 can also be used to identifyessentially derived varieties and other progeny varieties developed fromthe use of celery cultivar TBG 40, as well as cells and other plantparts thereof. Such plants may be developed using the markers identifiedin WO 00/31964, U.S. Pat. Nos. 6,162,967, and 7,288,386. Progeny plantsand plant parts produced using celery cultivar TBG 40 may be identifiedby having a molecular marker profile of at least 25%, 30%, 35%, 40%,45%, 50%, 55%, 60%, 65%, 70%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%,83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99%, or 99.5% genetic contribution from celery cultivar TBG40, as measured by either percent identity or percent similarity. Suchprogeny may be further characterized as being within a pedigree distanceof celery cultivar TBG 40, such as within 1, 2, 3, 4, or 5 or lesscross-pollinations to a celery plant other than celery cultivar TBG 40or a plant that has celery cultivar TBG 40 as a progenitor. Uniquemolecular profiles may be identified with other molecular tools such asSNPs and RFLPs.

While determining the genotypic profile of the plants described supra,several unique SSR profiles may also be identified which did not appearin either parent of such plant. Such unique SSR profiles may ariseduring the breeding process from recombination or mutation. Acombination of several unique alleles provides a means of identifying aplant variety, an F₁ progeny produced from such variety, and progenyproduced from such variety.

Molecular data from TBG 40 may be used in a plant breeding process.Nucleic acids may be isolated from a seed of TBG 40 or from a plant,plant part, or cell produced by growing a seed of TBG 40, or from a seedof TBG 40 with a gene conversion, or from a plant, plant part, or cellof TBG 40 with a gene conversion. One or more polymorphisms may beisolated from the nucleic acids. A plant having one or more of theidentified polymorphisms may be selected and used in a plant breedingmethod to produce another plant.

In another embodiment of the invention, the genetic complement of thecelery cultivar TBG 40 is provided. The phrase “genetic complement” isused to refer to the aggregate of nucleotide sequences, the expressionof which sequences defines the phenotype of, in the present case, acelery plant, or a cell or tissue of that plant. A genetic complementthus represents the genetic makeup of a cell, tissue or plant, and ahybrid genetic complement represents the genetic makeup of a hybridcell, tissue or plant. The invention thus provides celery plant cellsthat have a genetic complement in accordance with the celery plant cellsdisclosed herein, and plants, seeds and plants containing such cells.Plant genetic complements may be assessed by genetic marker profiles,and by the expression of phenotypic traits that are characteristic ofthe expression of the genetic complement, e.g., isozyme typing profiles.

Introduction of a New Trait or Locus into Celery Cultivar TBG 40

Cultivar TBG 40 represents a new base genetic variety into which a newgene, locus or trait may be introgressed. Backcrossing and directtransformation represent two important methods that can be used toaccomplish such an introgression.

Single Gene (Locus) Conversions

When the term “celery plant” is used in the context of the presentinvention, this also includes any single gene or locus conversions ofthat variety. The term “single locus converted plant” or “single geneconverted plant” refers to those celery plants which are developed bybackcrossing or genetic engineering, wherein essentially all of thedesired morphological and physiological characteristics of a variety arerecovered in addition to the one or more genes transferred into thevariety via the backcrossing technique or genetic engineering.Backcrossing methods can be used with the present invention to improveor introduce a characteristic into the variety.

A backcross conversion of celery cultivar TBG 40 occurs when DNAsequences are introduced through backcrossing (Hallauer, et al., “CornBreeding,” Corn and Corn Improvements, No. 18, pp. 463-481 (1988)), withcelery cultivar TBG 40 utilized as the recurrent parent. Both naturallyoccurring and transgenic DNA sequences may be introduced throughbackcrossing techniques. A backcross conversion may produce a plant witha trait or locus conversion in at least two or more backcrosses,including at least 2 crosses, at least 3 crosses, at least 4 crosses, atleast 5 crosses, and the like. Molecular marker assisted breeding orselection may be utilized to reduce the number of backcrosses necessaryto achieve the backcross conversion. For example, see, Openshaw, S. J.,et al., Marker-assisted Selection in Backcross Breeding, ProceedingsSymposium of the Analysis of Molecular Data, Crop Science Society ofAmerica, Corvallis, Oreg. (August 1994), where it is demonstrated that abackcross conversion can be made in as few as two backcrosses.

The complexity of the backcross conversion method depends on the type oftrait being transferred (single genes or closely linked genes ascompared to unlinked genes), the level of expression of the trait, thetype of inheritance (cytoplasmic or nuclear), and the types of parentsincluded in the cross. It is understood by those of ordinary skill inthe art that for single gene traits that are relatively easy toclassify, the backcross method is effective and relatively easy tomanage. (See, Hallauer, et al., Corn and Corn Improvement, Sprague andDudley, Third Ed. (1998)). Desired traits that may be transferredthrough backcross conversion include, but are not limited to, sterility(nuclear and cytoplasmic), fertility restoration, nutritionalenhancements, drought tolerance, nitrogen utilization, altered fattyacid profile, modified fatty acid metabolism, modified carbohydratemetabolism, industrial enhancements, yield stability, yield enhancement,disease resistance (bacterial, fungal, or viral), insect resistance, andherbicide resistance. In addition, an introgression site itself, such asan FRT site, Lox site, or other site specific integration site, may beinserted by backcrossing and utilized for direct insertion of one ormore genes of interest into a specific plant variety.

A single locus may contain several transgenes, such as a transgene fordisease resistance that, in the same expression vector, also contains atransgene for herbicide resistance. The gene for herbicide resistancemay be used as a selectable marker and/or as a phenotypic trait. Asingle locus conversion of site specific integration system allows forthe integration of multiple genes at a known recombination site in thegenome. At least one, at least two or at least three and less than ten,less than nine, less than eight, less than seven, less than six, lessthan five or less than four locus conversions may be introduced into theplant by backcrossing, introgression or transformation to express thedesired trait, while the plant, or a plant grown from the seed, plantpart or plant cell, otherwise retains the phenotypic characteristics ofthe deposited seed when grown under the same environmental conditions.

The backcross conversion may result from either the transfer of adominant allele or a recessive allele. Selection of progeny containingthe trait of interest is accomplished by direct selection for a traitassociated with a dominant allele. Transgenes transferred viabackcrossing typically function as a dominant single gene trait and arerelatively easy to classify. Selection of progeny for a trait that istransferred via a recessive allele requires growing and selfing thefirst backcross generation to determine which plants carry the recessivealleles. Recessive traits may require additional progeny testing insuccessive backcross generations to determine the presence of the locusof interest. The last backcross generation is usually selfed to givepure breeding progeny for the gene(s) being transferred, although abackcross conversion with a stably introgressed trait may also bemaintained by further backcrossing to the recurrent parent withselection for the converted trait.

Along with selection for the trait of interest, progeny are selected forthe phenotype of the recurrent parent. The backcross is a form ofinbreeding, and the features of the recurrent parent are automaticallyrecovered after successive backcrosses. Poehlman, Breeding Field Crops,p. 204 (1987). Poehlman suggests from one to four or more backcrosses,but as noted above, the number of backcrosses necessary can be reducedwith the use of molecular markers. Other factors, such as a geneticallysimilar donor parent, may also reduce the number of backcrossesnecessary. As noted by Poehlman, backcrossing is easiest for simplyinherited, dominant, and easily recognized traits.

One process for adding or modifying a trait or locus in celery cultivarTBG 40 comprises crossing celery cultivar TBG 40 plants grown fromcelery cultivar TBG 40 seed with plants of another celery variety thatcomprise the desired trait, gene or locus, selecting F₁ progeny plantsthat comprise the desired trait, gene or locus to produce selected F₁progeny plants, crossing the selected progeny plants with the celerycultivar TBG 40 plants to produce backcross progeny plants, selectingfor backcross progeny plants that have the desired trait, gene or locusand the morphological characteristics of celery cultivar TBG 40 toproduce selected backcross progeny plants, and backcrossing to celerycultivar TBG 40 three or more times in succession to produce selectedfourth or higher backcross progeny plants that comprise said trait, geneor locus. The modified celery cultivar TBG 40 may be furthercharacterized as having the physiological and morphologicalcharacteristics of celery cultivar TBG 40 as determined at the 5%significance level when grown in the same environmental conditionsand/or may be characterized by percent similarity or identity to celerycultivar TBG 40 as determined by SSR markers. The above method may beutilized with fewer backcrosses in appropriate situations, such as whenthe donor parent is highly related or markers are used in the selectionstep. Desired traits that may be used include those nucleic acids knownin the art, some of which are listed herein, that will affect traitsthrough nucleic acid expression or inhibition. Desired loci include theintrogression of FRT, Lox, and other sites for site specificintegration, which may also affect a desired trait if a functionalnucleic acid is inserted at the integration site.

In addition, the above process and other similar processes describedherein may be used to produce first generation progeny celery seed byadding a step at the end of the process that comprises crossing celerycultivar TBG 40 with the introgressed trait or locus with a differentcelery plant and harvesting the resultant first generation progenycelery seed.

Methods for Genetic Engineering of Celery

With the advent of molecular biological techniques that have allowed theisolation and characterization of genes that encode specific proteinproducts, scientists in the field of plant biology developed a stronginterest in engineering the genome of plants (genetic engineering) tocontain and express foreign genes, or additional, or modified versionsof native, or endogenous genes (perhaps driven by different promoters)in order to alter the traits of a plant in a specific manner. Plantsaltered by genetic engineering are often referred to as ‘geneticallymodified’. Any DNA sequences, whether from a different species or fromthe same species, which are introduced into the genome usingtransformation and/or various breeding methods, are referred to hereincollectively as “transgenes.” Over the last fifteen to twenty yearsseveral methods for producing transgenic plants have been developed, andthe present invention, in particular embodiments, also relates totransformed versions of the claimed cultivar.

Vectors used for the transformation of celery cells are not limited solong as the vector can express an inserted DNA in the cells. Forexample, vectors comprising promoters for constitutive gene expressionin celery cells (e.g., cauliflower mosaic virus 35S promoter) andpromoters inducible by exogenous stimuli can be used. Examples ofsuitable vectors include pBI binary vector. The “celery cell” into whichthe vector is to be introduced includes various forms of celery cells,such as cultured cell suspensions, protoplasts, leaf sections, andcallus. A vector can be introduced into celery cells by known methods,such as the polyethylene glycol method, polycation method,electroporation, Agrobacterium-mediated transfer, particle bombardmentand direct DNA uptake by protoplasts. See, e.g., Pang et al. (The PlantJ., 9, 899-909, 1996).

Numerous methods for plant transformation have been developed, includingbiological and physical, plant transformation protocols. See, forexample, Miki, et al., “Procedures for Introducing Foreign DNA intoPlants” in Methods in Plant Molecular Biology and Biotechnology, Glickand Thompson (Eds.), CRC Press, Inc., Boca Raton, pp. 67-88 (1993). Inaddition, expression vectors and in vitro culture methods for plant cellor tissue transformation and regeneration of plants are available. See,for example, Gruber, et al., “Vectors for Plant Transformation” inMethods in Plant Molecular Biology and Biotechnology, Glick and Thompson(Eds.), CRC Press, Inc., Boca Raton, pp. 89-119 (1993).

A. Agrobacterium-Mediated Transformation:

One method for introducing an expression vector into plants is based onthe natural transformation system of Agrobacterium. See, for example,Horsch, et al., Science, 227:1229 (1985). A. tumefaciens and A.rhizogenes are plant pathogenic soil bacteria which geneticallytransform plant cells. The Ti and Ri plasmids of A. tumefaciens and A.rhizogenes, respectively, carry genes responsible for genetictransformation of the plant. See, for example, Kado, C. I., Crit. Rev.Plant Sci., 10:1 (1991). Descriptions of Agrobacterium vector systemsand methods for Agrobacterium-mediated gene transfer are provided byGruber, et al., supra, Miki, et al., supra, and Moloney, et al., PlantCell Rep., 8:238 (1989). See also, U.S. Pat. No. 5,563,055 (Townsend andThomas), issued Oct. 8, 1996.

Agrobacterium-mediated transfer is a widely applicable system forintroducing gene loci into plant cells, including celery. An advantageof the technique is that DNA can be introduced into whole plant tissues,thereby bypassing the need for regeneration of an intact plant from aprotoplast. Modern Agrobacterium transformation vectors are capable ofreplication in E. coli as well as Agrobacterium, allowing for convenientmanipulations (Klee et al., Bio. Tech., 3(7):637-642, 1985). Moreover,recent technological advances in vectors for Agrobacterium-mediated genetransfer have improved the arrangement of genes and restriction sites inthe vectors to facilitate the construction of vectors capable ofexpressing various polypeptide coding genes. The vectors described haveconvenient multi-linker regions flanked by a promoter and apolyadenylation site for direct expression of inserted polypeptidecoding genes. Additionally, Agrobacterium containing both armed anddisarmed Ti genes can be used for transformation.

In those plant strains where Agrobacterium-mediated transformation isefficient, it is the method of choice because of the facile and definednature of the gene locus transfer. The use of Agrobacterium-mediatedplant integrating vectors to introduce DNA into plant cells is wellknown in the art (Fraley et al., Bio. Tech., 3(7):629-635, 1985; U.S.Pat. No. 5,563,055). For example, U.S. Pat. No. 5,349,124 describes amethod of transforming celery plant cells using Agrobacterium-mediatedtransformation. By inserting a chimeric gene having a DNA codingsequence encoding for the full-length B.t. toxin protein that expressesa protein toxic toward Lepidopteran larvae, this methodology resulted incelery having resistance to such insects.

B. Direct Gene Transfer:

Several methods of plant transformation, collectively referred to asdirect gene transfer, have been developed as an alternative toAgrobacterium-mediated transformation. A generally applicable method fordelivering transforming DNA segments to plant cells ismicroprojectile-mediated transformation, or microprojectile bombardment.In this method, particles are coated with nucleic acids and deliveredinto cells by a propelling force. Sanford, et al., Part. Sci. Technol.,5:27 (1987); Sanford, J. C., Trends Biotech., 6:299 (1988); Klein, etal., Bio/technology, 6:559-563 (1988); Sanford, J. C., Physiol Plant,7:206 (1990); Klein, et al., Bio/technology, 10:268 (1992). See also,U.S. Pat. No. 5,015,580 (Christou, et al.), issued May 14, 1991; U.S.Pat. No. 5,322,783 (Tomes, et al.), issued Jun. 21, 1994.

Another method for physical delivery of DNA to plants is sonication oftarget cells. Zhang, et al., Bio/technology, 9:996 (1991).Alternatively, liposome and spheroplast fusion have been used tointroduce expression vectors into plants. Deshayes, et al., EMBO J,4:2731 (1985); Christou, et al., PNAS, 84:3962 (1987). Direct uptake ofDNA into protoplasts using CaCl₂) precipitation, calcium phosphateprecipitation, polyethylene glycol treatment, polyvinyl alcohol, orpoly-L-ornithine has also been reported. See, e.g., Potrykus et al.,Mol. Gen. Genet., 199:183-188, 1985; Omirulleh et al., Plant Mol. Biol.,21(3):415-428, 1993; Fromm et al., Nature, 312:791-793, 1986; Uchimiyaet al., Mol. Gen. Genet., 204:204, 1986; Marcotte et al., Nature,335:454, 1988; Hain, et al., Mol. Gen. Genet., 199:161, 1985 and Draper,et al., Plant Cell Physiol. 23:451, 1982.

Electroporation of protoplasts and whole cells and tissues has also beendescribed. Donn, et al., In Abstracts of VIIth International Congress onPlant Cell and Tissue Culture IAPTC, A2-38, p. 53, 1990; D'Halluin, etal., Plant Cell, 4:1495-1505, 1992; and Spencer, et al., Plant Mol.Biol., 24:51-61, 1994. Another illustrative embodiment of a method fordelivering DNA into plant cells by acceleration is the BiolisticsParticle Delivery System, which can be used to propel particles coatedwith DNA or cells through a screen, such as a stainless steel or Nytexscreen, onto a surface covered with target celery cells.

Transformation of plants and expression of foreign genetic elements isexemplified in Choi et al., Plant Cell Rep., 13: 344-348, 1994 and Ellulet al., Theor. Appl. Genet., 107:462-469, 2003.

Following transformation of celery target tissues, expression ofselectable marker genes allows for preferential selection of transformedcells, tissues, and/or plants, using regeneration and selection methodsnow well known in the art.

The methods described herein for transformation would typically be usedfor producing a transgenic variety. The transgenic variety could then becrossed, with another (non-transformed or transformed) variety, in orderto produce a new transgenic variety. Alternatively, a genetic traitwhich has been engineered into a particular celery cultivar using thetransformation techniques described could be moved into another cultivarusing traditional backcrossing techniques that are well known in theplant breeding arts. For example, a backcrossing approach could be usedto move an engineered trait from a public, non-elite variety into anelite variety, or from a variety containing a foreign gene in its genomeinto a variety or varieties which do not contain that gene. As usedherein, “crossing” can refer to a simple X by Y cross, or the process ofbackcrossing, depending on the context.

Expression Vectors for Celery Transformation: Marker Genes

Expression vectors include at least one genetic marker, operably linkedto a regulatory element (a promoter, for example) that allowstransformed cells containing the marker to be either recovered bynegative selection, i.e., inhibiting growth of cells that do not containthe selectable marker gene, or by positive selection, i.e., screeningfor the product encoded by the genetic marker. Many commonly usedselectable marker genes for plant transformation are well known in thetransformation arts, and include, for example, genes that code forenzymes that metabolically detoxify a selective chemical agent which maybe an antibiotic or a herbicide, or genes that encode an altered targetwhich is insensitive to the inhibitor. A few positive selection methodsare also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) gene, isolated from transposonTn5, which when placed under the control of plant regulatory signalswhich confers resistance to kanamycin. Fraley et al., Proc. Natl. Acad.Sci. U.S.A., 80:4803 (1983). Another commonly used selectable markergene is the hygromycin phosphotransferase gene which confers resistanceto the antibiotic hygromycin. Vanden Elzen et al., Plant Mol. Biol.,5:299 (1985).

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase, aminoglycoside-3′-adenyl transferase,the bleomycin resistance determinant. Hayford et al., Plant Physiol.86:1216 (1988), Jones et al., Mol. Gen. Genet., 210:86 (1987), Svab etal., Plant Mol. Biol. 14:197 (1990) Hille et al., Plant Mol. Biol. 7:171(1986). Other selectable marker genes confer resistance to herbicidessuch as glyphosate, glufosinate or bromoxynil. Comai et al., Nature317:741-744 (1985), Gordon-Kamm et al., Plant Cell 2:603-618 (1990) andStalker et al., Science 242:419-423 (1988). Selectable marker genes forplant transformation not of bacterial origin include, for example, mousedihydrofolate reductase, plant 5-enolpyruvylshikimate-3-phosphatesynthase and plant acetolactate synthase. Eichholtz et al., Somatic CellMol. Genet. 13:67 (1987), Shah et al., Science 233:478 (1986), Charestet al., Plant Cell Rep. 8:643 (1990).

Another class of marker genes for plant transformation requiresscreening of presumptively transformed plant cells rather than directgenetic selection of transformed cells for resistance to a toxicsubstance such as an antibiotic. These genes are particularly useful toquantify or visualize the spatial pattern of expression of a gene inspecific tissues and are frequently referred to as reporter genesbecause they can be fused to a gene or gene regulatory sequence for theinvestigation of gene expression. Commonly used genes for screeningpresumptively transformed cells include α-glucuronidase (GUS,α-galactosidase, luciferase and chloramphenicol, acetyltransferase.Jefferson, R. A., Plant Mol. Biol. Rep. 5:387 (1987), Teeri et al., EMBOJ. 8:343 (1989), Koncz et al., Proc. Natl. Acad. Sci U.S.A. 84:131(1987), DeBlock et al., EMBO J. 3:1681 (1984).

In vivo methods for visualizing GUS activity that do not requiredestruction of plant tissue are available. Molecular Probes publication2908, IMAGENE GREEN, p. 1-4 (1993) and Naleway et al., J. Cell Biol.115:151a (1991). However, these in vivo methods for visualizing GUSactivity have not proven useful for recovery of transformed cellsbecause of low sensitivity, high fluorescent backgrounds and limitationsassociated with the use of luciferase genes as selectable markers.

More recently, a gene encoding Green Fluorescent Protein (GFP) has beenutilized as a marker for gene expression in prokaryotic and eukaryoticcells. Chalfie et al., Science 263:802 (1994). GFP and mutants of GFPmay be used as screenable markers.

Expression Vectors for Celery Transformation: Promoters

Genes included in expression vectors must be driven by a nucleotidesequence comprising a regulatory element, for example, a promoter.Several types of promoters are now well known in the transformationarts, as are other regulatory elements that can be used alone or incombination with promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Examples of promoters under developmental control includepromoters that preferentially initiate transcription in certain tissues,such as leaves, roots, seeds, fibers, xylem vessels, tracheids, orsclerenchyma. Such promoters are referred to as “tissue-preferred”.Promoters which initiate transcription only in certain tissue arereferred to as “tissue-specific”. A “cell type” specific promoterprimarily drives expression in certain cell types in one or more organs,for example, vascular cells in roots or leaves. An “inducible” promoteris a promoter which is under environmental control. Examples ofenvironmental conditions that may effect transcription by induciblepromoters include anaerobic conditions or the presence of light.Tissue-specific, tissue-preferred, cell type specific, and induciblepromoters constitute the class of “non-constitutive” promoters. A“constitutive” promoter is a promoter which is active under mostenvironmental conditions.

A. Inducible Promoters:

An inducible promoter is operably linked to a gene for expression incelery. Optionally, the inducible promoter is operably linked to anucleotide sequence encoding a signal sequence which is operably linkedto a gene for expression in celery. With an inducible promoter the rateof transcription increases in response to an inducing agent.

Any inducible promoter can be used in the instant invention. See Ward etal., Plant Mol. Biol. 22:361-366 (1993). Exemplary inducible promotersinclude, but are not limited to, that from the ACEI system whichresponds to copper (Meft et al., PNAS 90:4567-4571 (1993)); In2 genefrom maize which responds to benzenesulfonamide herbicide safeners(Hershey et al., Mol. Gen Genetics 227:229-237 (1991) and Gatz et al.,Mol. Gen. Genetics 243:32-38 (1994)) or Tet repressor from Tn10 (Gatz etal., Mol. Gen. Genetics 227:229-237 (1991). A particularly preferredinducible promoter is a promoter that responds to an inducing agent towhich plants do not normally respond. An exemplary inducible promoter isthe inducible promoter from a steroid hormone gene, the transcriptionalactivity of which is induced by a glucocorticosteroid hormone. Schena etal., Proc. Natl. Acad. Sci. U.S.A. 88:0421 (1991).

B. Constitutive Promoters:

A constitutive promoter is operably linked to a gene for expression incelery or the constitutive promoter is operably linked to a nucleotidesequence encoding a signal sequence which is operably linked to a genefor expression in celery.

Many different constitutive promoters can be utilized in the instantinvention. Exemplary constitutive promoters include, but are not limitedto, the promoters from plant viruses such as the 35S promoter from CaMV(Odell et al., Nature 313:810-812 (1985) and the promoters from suchgenes as rice actin (McElroy et al., Plant Cell 2:163-171 (1990));ubiquitin (Christensen et al., Plant Mol. Biol. 12:619-632 (1989) andChristensen et al., Plant Mol. Biol. 18:675-689 (1992)); pEMU (Last etal., Theor. Appl. Genet. 81:581-588 (1991)); MAS (Velten et al., EMBO J.3:2723-2730 (1984)) and maize H3 histone (Lepetit et al., Mol. Gen.Genetics 231:276-285 (1992) and Atanassova et al., Plant Journal 2 (3):291-300 (1992)). The ALS promoter, Xbal/Ncol fragment 5′ to the Brassicanapus ALS3 structural gene (or a nucleotide sequence similarity to saidXbal/Ncol fragment), represents a particularly useful constitutivepromoter. See PCT application WO 96/30530.

C. Tissue-Specific or Tissue-Preferred Promoters:

A tissue-specific promoter is operably linked to a gene for expressionin celery. Optionally, the tissue-specific promoter is operably linkedto a nucleotide sequence encoding a signal sequence which is operablylinked to a gene for expression in celery. Plants transformed with agene of interest operably linked to a tissue-specific promoter producethe protein product of the transgene exclusively, or preferentially, ina specific tissue.

Any tissue-specific or tissue-preferred promoter can be utilized in theinstant invention. Exemplary tissue-specific or tissue-preferredpromoters include, but are not limited to, a root-preferred promoter,such as that from the phaseolin gene (Murai et al., Science 23:476-482(1983) and Sengupta-Gopalan et al., Proc. Natl. Acad. Sci. U.S.A.82:3320-3324 (1985)); a leaf-specific and light-induced promoter such asthat from cab or rubisco (Simpson et al., EMBO J. 4(11):2723-2729 (1985)and Timko et al., Nature 318:579-582 (1985)); an anther-specificpromoter such as that from LAT52 (Twell et al., Mol. Gen. Genetics217:240-245 (1989)); a pollen-specific promoter such as that from Zm13(Guerrero et al., Mol. Gen. Genetics 244:161-168 (1993)) or amicrospore-preferred promoter such as that from apg (Twell et al., Sex.Plant Reprod. 6:217-224 (1993)).

Signal Sequences for Targeting Proteins to Subcellular Compartments

Transport of protein produced by transgenes to a subcellular compartmentsuch as the chloroplast, vacuole, peroxisome, glyoxysome, cell wall ormitochondrion or for secretion into the apoplast, is accomplished bymeans of operably linking the nucleotide sequence encoding a signalsequence to the 5′ and/or 3′ region of a gene encoding the protein ofinterest. Targeting sequences at the 5′ and/or 3′ end of the structuralgene may determine, during protein synthesis and processing, where theencoded protein is ultimately compartmentalized.

The presence of a signal sequence directs a polypeptide to either anintracellular organelle or subcellular compartment or for secretion tothe apoplast. Many signal sequences are known in the art. See, forexample Becker et al., Plant Mol. Biol. 20:49 (1992), Close, P. S.,Master's Thesis, Iowa State University (1993), Knox, C., et al.,“Structure and Organization of Two Divergent Alpha-Amylase Genes fromBarley”, Plant Mol. Biol. 9:3-17 (1987), Lerner et al., Plant Physiol.91:124-129 (1989), Fontes et al., Plant Cell 3:483-496 (1991), Matsuokaet al., Proc. Natl. Acad. Sci. 88:834 (1991), Gould et al., J. Cell.Biol. 108:1657 (1989), Creissen et al., Plant J 2:129 (1991), Kalderon,et al., A short amino acid sequence able to specify nuclear location,Cell 39:499-509 (1984), Steifel, et al., Expression of a maize cell wallhydroxyproline-rich glycoprotein gene in early leaf and root vasculardifferentiation, Plant Cell 2:785-793 (1990).

Additional Methods for Genetic Engineering of Celery

In general, methods to transform, modify, edit or alter plant endogenousgenomic DNA include altering the plant native DNA sequence or apre-existing transgenic sequence including regulatory elements, codingand non-coding sequences. These methods can be used, for example, totarget nucleic acids to pre-engineered target recognition sequences inthe genome. Such pre-engineered target sequences may be introduced bygenome editing or modification. As an example, a genetically modifiedplant variety is generated using “custom” or engineered endonucleasessuch as meganucleases produced to modify plant genomes (see e.g., WO2009/114321; Gao et al. (2010) Plant Journal 1:176-187). Anothersite-directed engineering method is through the use of zinc fingerdomain recognition coupled with the restriction properties ofrestriction enzyme. See e.g., Urnov, et al., (2010) Nat Rev Genet.11(9):636-46; Shukla, et al., (2009) Nature 459 (7245):437-41. Atranscription activator-like (TAL) effector-DNA modifying enzyme (TALEor TALEN) is also used to engineer changes in plant genome. See e.g.,US20110145940, Cermak et al., (2011) Nucleic Acids Res. 39(12) and Bochet al., (2009), Science 326(5959): 1509-12. Site-specific modificationof plant genomes can also be performed using the bacterial type IICRISPR (clustered regularly interspaced short palindromic repeats)/Cas(CRISPR-associated) system, as well as similar CRISPR relatedtechnologies. See e.g., Belhaj et al., (2013), Plant Methods 9: 39; TheCas9/guide RNA-based system allows targeted cleavage of genomic DNAguided by a customizable small noncoding RNA in plants (see e.g., WO2015026883A1, incorporated herein by reference).

A genetic map can be generated that identifies the approximatechromosomal location of an integrated DNA molecule, for example viaconventional restriction fragment length polymorphisms (RFLP),polymerase chain reaction (PCR) analysis, simple sequence repeats (SSR),and single nucleotide polymorphisms (SNP). For exemplary methodologiesin this regard, see Glick and Thompson, Methods in Plant MolecularBiology and Biotechnology, pp. 269-284 (CRC Press, Boca Raton, 1993).

Wang et al. discuss “Large Scale Identification, Mapping and Genotypingof Single-Nucleotide Polymorphisms in the Human Genome”, Science (1998)280:1077-1082, and similar capabilities are increasingly available forthe celery genome. Map information concerning chromosomal location isuseful for proprietary protection of a subject transgenic plant. Ifunauthorized propagation is undertaken and crosses made with othergermplasm, the map of the integration region can be compared to similarmaps for suspect plants to determine if the latter have a commonparentage with the subject plant. Map comparisons could involvehybridizations, RFLP, PCR, SSR, sequencing or combinations thereof, allof which are conventional techniques. SNPs may also be used alone or incombination with other techniques.

Celery Cultivar TBG 40 Further Comprising a Transgene

Transgenes and transformation methods provide means to engineer thegenome of plants to contain and express heterologous genetic elements,including but not limited to foreign genetic elements, additional copiesof endogenous elements, and/or modified versions of native or endogenousgenetic elements, in order to alter at least one trait of a plant in aspecific manner. Any heterologous DNA sequence(s), whether from adifferent species or from the same species, which are inserted into thegenome using transformation, backcrossing, or other methods known to oneof skill in the art are referred to herein collectively as transgenes.The sequences are heterologous based on sequence source, location ofintegration, operably linked elements, or any combination thereof. Oneor more transgenes of interest can be introduced into celery cultivarTBG 40. Transgenic variants of celery cultivar TBG 40 plants, seeds,cells, and parts thereof or derived therefrom are provided. Transgenicvariants of TBG 40 comprise the physiological and morphologicalcharacteristics of celery cultivar TBG 40, such as determined at the 5%significance level when grown in the same environmental conditions,and/or may be characterized or identified by percent similarity oridentity to TBG 40 as determined by SSR or other molecular markers. Insome examples, transgenic variants of celery cultivar TBG 40 areproduced by introducing at least one transgene of interest into celerycultivar TBG 40 by transforming TBG 40 with a polynucleotide comprisingthe transgene of interest. In other examples, transgenic variants ofcelery cultivar TBG 40 are produced by introducing at least onetransgene by introgressing the transgene into celery cultivar TBG 40 bycrossing.

In one example, a process for modifying celery cultivar TBG 40 with theaddition of a desired trait, said process comprising transforming acelery plant of cultivar TBG 40 with a transgene that confers a desiredtrait is provided. Therefore, transgenic TBG 40 celery cells, plants,plant parts, and seeds produced from this process are provided. In someexamples one more desired traits may include traits such as sterility(nuclear and cytoplasmic), fertility restoration, nutritionalenhancements, drought tolerance, nitrogen utilization, altered fattyacid profile, modified fatty acid metabolism, modified carbohydratemetabolism, industrial enhancements, yield stability, yield enhancement,disease resistance (bacterial, fungal, or viral), insect resistance, andherbicide resistance. The specific gene may be any known in the art orlisted herein, including but not limited to a polynucleotide conferringresistance to an ALS-inhibitor herbicide, imidazolinone, sulfonylurea,protoporphyrinogen oxidase (PPO) inhibitors, hydroxyphenyl pyruvatedioxygenase (HPPD) inhibitors, glyphosate, glufosinate, triazine,2,4-dichlorophenoxyacetic acid (2,4-D), dicamba, broxynil, metribuzin,or benzonitrile herbicides; a polynucleotide encoding a Bacillusthuringiensis polypeptide, a polynucleotide encoding a phytase, a fattyacid desaturase (e.g., FAD-2, FAD-3), galactinol synthase, a raffinosesynthetic enzyme; or a polynucleotide conferring resistance to tipburn,Fusarium oxysporum, Nasonovia ribisnigri, Sclerotinia sclerotiorum orother plant pathogens.

Foreign Protein Genes and Agronomic Genes

By means of the present invention, plants can be genetically engineeredto express various phenotypes of agronomic interest. Through thetransformation of celery, the expression of genes can be altered toenhance disease resistance, insect resistance, herbicide resistance,agronomic, nutritional quality, and other traits. Transformation canalso be used to insert DNA sequences which control or help controlmale-sterility. DNA sequences native to celery, as well as non-nativeDNA sequences, can be transformed into celery and used to alter levelsof native or non-native proteins. Various promoters, targetingsequences, enhancing sequences, and other DNA sequences can be insertedinto the genome for the purpose of altering the expression of proteins.Reduction of the activity of specific genes (also known as genesilencing or gene suppression) is desirable for several aspects ofgenetic engineering in plants.

Many techniques for gene silencing are well known to one of skill in theart, including, but not limited to, knock-outs (such as by insertion ofa transposable element such as mu (Vicki Chandler, The Maize Handbook,Ch. 118 (Springer-Verlag 1994)) or other genetic elements such as a FRTand Lox that are used for site specific integrations, antisensetechnology (see, e.g., Sheehy, et al., PNAS USA, 85:8805-8809 (1988);and U.S. Pat. Nos. 5,107,065, 5,453,566, and 5,759,829); co-suppression(e.g., Taylor, Plant Cell, 9:1245 (1997); Jorgensen, Trends Biotech.,8(12):340-344 (1990); Flavell, PNAS USA, 91:3490-3496 (1994); Finnegan,et al., Bio/Technology, 12:883-888 (1994); Neuhuber, et al., Mol. Gen.Genet., 244:230-241 (1994)); RNA interference (Napoli, et al., PlantCell, 2:279-289 (1990); U.S. Pat. No. 5,034,323; Sharp, Genes Dev.,13:139-141 (1999); Zamore, et al., Cell, 101:25-33 (2000); Montgomery,et al., PNAS USA, 95:15502-15507 (1998)), virus-induced gene silencing(Burton, et al., Plant Cell, 12:691-705 (2000); Baulcombe, Curr. Op.Plant Bio., 2:109-113 (1999)); target-RNA-specific ribozymes (Haseloff,et al., Nature, 334: 585-591 (1988)); hairpin structures (Smith, et al.,Nature, 407:319-320 (2000); WO 99/53050; WO 98/53083); MicroRNA(Aukerman & Sakai, Plant Cell, 15:2730-2741 (2003)); ribozymes(Steinecke, et al., EMBO J., 11:1525 (1992); Perriman, et al., AntisenseRes. Dev., 3:253 (1993)); oligonucleotide mediated targeted modification(e.g., WO 03/076574 and WO 99/25853); Zn-finger targeted molecules(e.g., WO 01/52620, WO 03/048345, and WO 00/42219); and other methods orcombinations of the above methods known to those of skill in the art.

Likewise, by means of the present invention, agronomic genes can beexpressed in transformed plants. More particularly, plants can begenetically engineered to express various phenotypes of agronomicinterest. Exemplary nucleotide sequences and/or native loci that conferat least one trait of interest, which optionally may be conferred oraltered by genetic engineering, transformation or introgression of atransformed event include, but are not limited to, those categorizedbelow:

1. Genes that Confer Resistance to Pests or Disease and that Encode:

A. Plant disease resistance genes. Plant defenses are often activated byspecific interaction between the product of a disease resistance gene(R) in the plant and the product of a corresponding avirulence (Avr)gene in the pathogen. A plant line can be transformed with a clonedresistance gene to engineer plants that are resistant to specificpathogen strains. See, for example Jones et al., Science 266:789 (1994)(cloning of the tomato Cf-9 gene for resistance to Cladosporium fulvum);Martin et al., Science 262:1432 (1993) (tomato Pto gene for resistanceto Pseudomonas syringae pv. Tomato encodes a protein kinase); Mindrinoset al., Cell 78:1089 (1994) (Arabidopsis RSP2 gene for resistance toPseudomonas syringae).

B. A Bacillus thuringiensis protein, a derivative thereof or a syntheticpolypeptide modeled thereon. See, for example, Geiser et al., Gene48:109 (1986), who disclose the cloning and nucleotide sequence of a Btδ-endotoxin gene. Moreover, DNA molecules encoding δ-endotoxin genes canbe purchased from American Type Culture Collection, Manassas, Va., forexample, under ATCC Accession Nos. 40098, 67136, 31995 and 31998.

C. A lectin. See, for example, the disclosure by Van Damme et al., PlantMolec. Biol. 24:25 (1994), who disclose the nucleotide sequences ofseveral Clivia miniata mannose-binding lectin genes.

D. A vitamin-binding protein such as avidin. See PCT applicationUS93/06487, the contents of which are hereby incorporated by reference.The application teaches the use of avidin and avidin homologues aslarvicides against insect pests.

E. An enzyme inhibitor, for example, a protease or proteinase inhibitoror an amylase inhibitor. See, for example, Abe et al., J. Biol. Chem.262:16793 (1987) (nucleotide sequence of rice cysteine proteinaseinhibitor), Huub et al., Plant Molec. Biol. 21:985 (1993) (nucleotidesequence of cDNA encoding tobacco proteinase inhibitor I), Sumitani etal., Biosci. Biotech. Biochem. 57:1243 (1993) (nucleotide sequence ofStreptomyces nitrosporeus α-amylase inhibitor).

F. An insect-specific hormone or pheromone such as an ecdysteroid andjuvenile hormone, a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof. See, for example, the disclosure byHammock et al., Nature 344:458 (1990), of baculovirus expression ofcloned juvenile hormone esterase, an inactivator of juvenile hormone.

G. An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. For example, see thedisclosures of Regan, J. Biol. Chem. 269:9 (1994) (expression cloningyields DNA coding for insect diuretic hormone receptor), and Pratt etal., Biochem. Biophys. Res. Comm. 163:1243 (1989) (an allostatin isidentified in Diploptera puntata). See also U.S. Pat. No. 5,266,317 toTomalski et al., who disclose genes encoding insect-specific, paralyticneurotoxins.

H. An insect-specific venom produced in nature by a snake, a wasp, etc.For example, see Pang et al., Gene 116:165 (1992), for disclosure ofheterologous expression in plants of a gene coding for a scorpioninsectotoxic peptide.

I. An enzyme responsible for a hyper accumulation of a monoterpene, asesquiterpene, a steroid, hydroxamic acid, a phenylpropanoid derivativeor another non-protein molecule with insecticidal activity.

J. An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, a glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme,a nuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase and a glucanase, whether natural or synthetic. See PCTapplication WO 93/02197 in the name of Scott et al., which discloses thenucleotide sequence of a callase gene. DNA molecules which containchitinase-encoding sequences can be obtained, for example, from ATCCunder Accession Nos. 39637 and 67152. See also Kramer et al., InsectBiochem. Molec. Biol. 23:691 (1993), who teach the nucleotide sequenceof a cDNA encoding tobacco hookworm chitinase, and Kawalleck et al.,Plant Molec. Biol. 21:673 (1993), who provide the nucleotide sequence ofthe parsley ubi4-2 polyubiquitin gene.

K. A molecule that stimulates signal transduction. For example, see thedisclosure by Botella et al., Plant Molec. Biol. 24:757 (1994), ofnucleotide sequences for mung bean calmodulin cDNA clones, and Griess etal., Plant Physiol. 104:1467 (1994), who provide the nucleotide sequenceof a maize calmodulin cDNA clone.

L. A hydrophobic moment peptide. See PCT application WO 95/16776(disclosure of peptide derivatives of Tachyplesin which inhibit fungalplant pathogens) and PCT application WO 95/18855 (teaches syntheticantimicrobial peptides that confer disease resistance), the respectivecontents of which are hereby incorporated by reference.

M. A membrane permease, a channel former or a channel blocker. Forexample, see the disclosure of Jaynes et al., Plant Sci 89:43 (1993), ofheterologous expression of a cecropin-β, lytic peptide analog to rendertransgenic tobacco plants resistant to Pseudomonas solanacearum.

N. A viral-invasive protein or a complex toxin derived therefrom. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. See Beachy et al., Ann. rev. Phytopathol.28:451 (1990). Coat protein-mediated resistance has been conferred upontransformed plants against alfalfa mosaic virus, cucumber mosaic virus,tobacco streak virus, potato virus X, potato virus Y, tobacco etchvirus, tobacco rattle virus and tobacco mosaic virus. Id.

O. An insect-specific antibody or an immunotoxin derived therefrom.Thus, an antibody targeted to a critical metabolic function in theinsect gut would inactivate an affected enzyme, killing the insect. CfTaylor et al., Abstract #497, Seventh Int'l Symposium on MolecularPlant-Microbe Interactions (Edinburgh, Scotland) (1994) (enzymaticinactivation in transgenic tobacco via production of single-chainantibody fragments).

P. A virus-specific antibody. See, for example, Tavladoraki et al.,Nature 366:469 (1993), who show that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

Q. A developmental-arrestive protein produced in nature by a pathogen ora parasite. Thus, fungal endo α-1, 4-D-polygalacturonases facilitatefungal colonization and plant nutrient release by solubilizing plantcell wall homo-α-1,4-D-galacturonase. See Lamb et al., Bio/Technology10:1436 (1992). The cloning and characterization of a gene which encodesa celery endopolygalacturonase-inhibiting protein is described byToubart et al., Plant J. 2:367 (1992).

R. A developmental-arrestive protein produced in nature by a plant. Forexample, Logemann et al., Bio/Technology 10:305 (1992), have shown thattransgenic plants expressing the barley ribosome-inactivating gene havean increased resistance to fungal disease.

S. A lettuce mosaic potyvirus (LMV) coat protein gene introduced intocelery in order to increase its resistance to LMV infection. See Dinantet al., Molecular Breeding. 1997, 3: 1, 75-86.

T. Genes involved in the Systemic Acquired Resistance (SAR) Responseand/or the pathogenesis-related genes. Briggs, S., Current Biology, 5(2)(1995).

U. Antifungal genes. See Cornelissen and Melchers, Plant Physiol.,101:709-712 (1993); Parijs et al., Planta 183:258-264 (1991) andBushnell et al., Can. J. of Plant Path. 20(2):137-149 (1998).

V. Genes that confer resistance to Phytophthora root rot, such as theRps 1, Rps 1-a, Rps 1-b, Rps 1-c, Rps 1-d, Rps 1-e, Rps 1-k, Rps 2, Rps3-a, Rps 3-b, Rps 3-c, Rps 4, Rps 5, Rps 6, Rps 7 and other Rps genes.See, for example, Shoemaker et al., Phytophthora Root Rot ResistanceGene Mapping in Soybean, Plant Genome IV Conference, San Diego, Calif.(1995).

2. Genes that Confer Resistance to an Herbicide:

A. An herbicide that inhibits the growing point or meristem, such as animidazolinone or a sulfonylurea. Exemplary genes in this category codefor mutant ALS and AHAS enzyme as described, for example, by Lee et al.,EMBO J. 7:1241 (1988), and Miki et al., Theor. Appl. Genet. 80:449(1990), respectively.

B. Glyphosate (resistance impaired by mutant5-enolpyruvlshikimate-3-phosphate synthase (EPSPS) and aroA genes,respectively) and other phosphono compounds such as glufosinate(phosphinothricin acetyl transferase, PAT and Streptomyces hygroscopicusphosphinothricin-acetyl transferase, bar, genes), and pyridinoxy orphenoxy proprionic acids and cyclohexones (ACCase inhibitor-encodinggenes). See, for example, U.S. Pat. No. 4,940,835 to Shah, et al., whichdiscloses the nucleotide sequence of a form of EPSPs which can conferglyphosate resistance. A DNA molecule encoding a mutant aroA gene can beobtained under ATCC accession number 39256, and the nucleotide sequenceof the mutant gene is disclosed in U.S. Pat. No. 4,769,061 to Comai. Seealso Umaballava-Mobapathie in Transgenic Research. 1999, 8: 1, 33-44that discloses Lactuca sativa resistant to glufosinate. European patentapplication No. 0 333 033 to Kumada et al., and U.S. Pat. No. 4,975,374to Goodman et al., disclose nucleotide sequences of glutamine synthetasegenes which confer resistance to herbicides such as L-phosphinothricin.The nucleotide sequence of a phosphinothricin-acetyl-transferase gene isprovided in European application No. 0 242 246 to Leemans et al.,DeGreef et al., Bio/Technology 7:61 (1989), describe the production oftransgenic plants that express chimeric bar genes coding forphosphinothricin acetyl transferase activity. Exemplary of genesconferring resistance to phenoxy proprionic acids and cyclohexones, suchas sethoxydim and haloxyfop are the Accl-S1, Accl-S2 and Accl-S3 genesdescribed by Marshall et al., Theor. Appl. Genet. 83:435 (1992).

C. An herbicide that inhibits photosynthesis, such as a triazine (psbAand gs+ genes) and a benzonitrile (nitrilase gene). Przibilla et al.,Plant Cell 3:169 (1991), describe the transformation of Chlamydomonaswith plasmids encoding mutant psbA genes. Nucleotide sequences fornitrilase genes are disclosed in U.S. Pat. No. 4,810,648 to Stalker, andDNA molecules containing these genes are available under ATCC AccessionNos. 53435, 67441, and 67442. Cloning and expression of DNA coding for aglutathione S-transferase is described by Hayes et al., Biochem. J.285:173 (1992).

D. Acetohydroxy acid synthase, which has been found to make plants thatexpress this enzyme resistant to multiple types of herbicides, has beenintroduced into a variety of plants. See Hattori et al., Mol. Gen.Genet. 246:419, 1995. Other genes that confer tolerance to herbicidesinclude a gene encoding a chimeric protein of rat cytochrome P4507A1 andyeast NADPH-cytochrome P450 oxidoreductase (Shiota et al., PlantPhysiol., 106:17, 1994), genes for glutathione reductase and superoxidedismutase (Aono et al., Plant Cell Physiol. 36:1687, 1995), and genesfor various phosphotransferases (Datta et al., Plant Mol. Biol. 20:619,1992).

E. Protoporphyrinogen oxidase (PPO; protox) is the target of thePPO-inhibitor class of herbicides; a PPO-inhibitor resistant PPO genewas recently identified in Amaranthus tuberculatus (Patzoldt et al.,PNAS, 103(33):12329-2334, 2006). PPO is necessary for the production ofchlorophyll, which is necessary for all plant survival. The protoxenzyme serves as the target for a variety of herbicidal compounds. Theseherbicides also inhibit growth of all the different species of plantspresent, causing their total destruction. The development of plantscontaining altered protox activity which are resistant to theseherbicides are described in U.S. Pat. Nos. 6,288,306, 6,282,837,5,767,373, and International Publication WO 01/12825.

F. Genes that confer resistance to auxin or synthetic auxin herbicides.For example an aryloxyalkanoate dioxygenase (AAD) gene may conferresistance to arlyoxyalkanoate herbicides, such as 2,4-D, as well aspyridyloxyacetate herbicides, such as described in U.S. Pat. No.8,283,522, and US2013/0035233. In other examples, a dicambamonooxygenase (DMO) is used to confer resistance to dicamba. Otherpolynucleotides of interest related to auxin herbicides and/or usesthereof include, for example, the descriptions found in U.S. Pat. Nos.8,119,380; 7,812,224; 7,884,262; 7,855,326; 7,939,721; 7,105,724;7,022,896; 8,207,092; US2011/067134; and US2010/0279866. Any of theabove listed herbicide genes (1-6) can be introduced into the claimedcelery cultivar through a variety of means including, but not limitedto, transformation and crossing.

3. Genes that Confer or Contribute to a Value-Added Trait, Such as:

A. Increased iron content of the celery, for example by transforming aplant with a soybean ferritin gene as described in Goto et al., ActaHorticulturae. 2000, 521, 101-109.

B. Decreased nitrate content of leaves, for example by transforming acelery with a gene coding for a nitrate reductase. See for exampleCurtis et al., Plant Cell Report. 1999, 18:11, 889-896.

C. Increased sweetness of the celery by transferring a gene coding formonellin that elicits a flavor 100,000 times sweeter than sugar on amolar basis. See Penarrubia et al., Biotechnology. 1992, 10:5, 561-564.

D. Modified fatty acid metabolism, for example, by transforming a plantwith an antisense gene of stearyl-ACP desaturase to increase stearicacid content of the plant. See Knultzon et al., Proc. Natl. Acad. Sci.USA 89:2625 (1992).

E. Modified carbohydrate composition effected, for example, bytransforming plants with a gene coding for an enzyme that alters thebranching pattern of starch. See Shiroza et al., J. Bacteriol. 170:810(1988) (nucleotide sequence of Streptococcus mutantsfructosyltransferase gene), Steinmetz et al., Mol. Gen. Genet. 20:220(1985) (nucleotide sequence of Bacillus subtilis levansucrase gene), Penet al., Bio/Technology 10:292 (1992) (production of transgenic plantsthat express Bacillus lichenifonnis α-amylase), Elliot et al., PlantMolec. Biol. 21:515 (1993) (nucleotide sequences of tomato invertasegenes), Søgaard et al., J. Biol. Chem. 268:22480 (1993) (site-directedmutagenesis of barley α-amylase gene), and Fisher et al., Plant Physiol.102:1045 (1993) (maize endosperm starch branching enzyme II).

F. Altered antioxidant content or composition, such as alteration oftocopherol or tocotrienols. See, for example, U.S. Pat. Nos. 6,787,683,7,154,029, WO 00/68393 (involving the manipulation of antioxidant levelsthrough alteration of a phytl prenyl transferase (ppt)); WO 03/082899(through alteration of a homogentisate geranyl geranyl transferase(hggt)).

G. Modified bolting tolerance in plants for example, by transferring agene encoding for gibberellin 2-oxidase (U.S. Pat. No. 7,262,340).Bolting has also been modified using non-transformation methods; seeWittwer, S. H., et al. (1957) Science. 126(3262): 30-31; Booij, R. etal., (1995) Scientia Horticulturae. 63:143-154; and Booij, R. et al.,(1994) Scientia Horticulturae. 58:271-282.

H. Decreased browning of the celery, for example by transforming a plantwith an siRNA, RNAi or microRNA vector, or other suppression sequencecoding for polyphenol oxidase (PPO) to silence the expression of PPOgenes. See Araji et al. (2014) Plant Physiology 164:1191-1203, Chi etal. (2014) BMC Plant Biology 14:62, and Carter, N., (2012) Petition forDetermination of Nonregulated Status: Arctic™ Apple (Malus x domestica)Events GD743 and GS784, received by APHIS.

4. Genes that Control Male-Sterility

A. Introduction of a deacetylase gene under the control of atapetum-specific promoter and with the application of the chemicalN-Ac-PPT. See international publication WO 01/29237.

B. Introduction of various stamen-specific promoters. See internationalpublications WO 92/13956 and WO 92/13957.

C. Introduction of the barnase and the barstar genes. See Paul et al.,Plant Mol. Biol. 19:611-622, 1992).

For additional examples of nuclear male and female sterility systems andgenes, see also, U.S. Pat. Nos. 5,859,341, 6,297,426, 5,478,369,5,824,524, 5,850,014, and 6,265,640, all of which are herebyincorporated by reference.

5. Genes that Affect Abiotic Stress Resistance

Genes that affect abiotic stress resistance (including but not limitedto flowering, seed development, enhancement of nitrogen utilizationefficiency, altered nitrogen responsiveness, drought resistance ortolerance, cold resistance or tolerance, high or low light intensity,and salt resistance or tolerance) and increased yield under stress. Forexample, see: WO 00/73475 where water use efficiency is altered throughalteration of malate; U.S. Pat. Nos. 5,892,009, 5,965,705, 5,929,305,5,891,859, 6,417,428, 6,664,446, 6,706,866, 6,717,034, 6,801,104, WO2000/060089, WO 2001/026459, WO 2001/035725, WO 2001/034726, WO2001/035727, WO 2001/036444, WO 2001/036597, WO 2001/036598, WO2002/015675, WO 2002/017430, WO 2002/077185, WO 2002/079403, WO2003/013227, WO 2003/013228, WO 2003/014327, WO 2004/031349, WO2004/076638, WO 98/09521, and WO 99/38977 describing genes, includingCBF genes and transcription factors effective in mitigating the negativeeffects of freezing, high salinity, and drought on plants, as well asconferring other positive effects on plant phenotype; U.S. Publ. No.2004/0148654 and WO 01/36596, where abscisic acid is altered in plantsresulting in improved plant phenotype, such as increased yield and/orincreased tolerance to abiotic stress; WO 2000/006341, WO 04/090143,U.S. Pat. Nos. 7,531,723 and 6,992,237, where cytokinin expression ismodified resulting in plants with increased stress tolerance, such asdrought tolerance, and/or increased yield. See also, WO 02/02776, WO2003/052063, JP 2002281975, U.S. Pat. No. 6,084,153, WO 01/64898, andU.S. Pat. Nos. 6,177,275 and 6,107,547 (enhancement of nitrogenutilization and altered nitrogen responsiveness). For ethylenealteration, see, U.S. Publ. Nos. 2004/0128719, 2003/0166197, and WO2000/32761. For plant transcription factors or transcriptionalregulators of abiotic stress, see, e.g., U.S. Publ. Nos. 2004/0098764 or2004/0078852.

Other genes and transcription factors that affect plant growth andagronomic traits, such as yield, flowering, plant growth, and/or plantstructure, can be introduced or introgressed into plants. See, e.g., WO97/49811 (LHY), WO 98/56918 (ESD4), WO 97/10339, U.S. Pat. No. 6,573,430(TFL), U.S. Pat. No. 6,713,663 (FT), U.S. Pat. Nos. 6,794,560, 6,307,126(GAI), WO 96/14414 (CON), WO 96/38560, WO 01/21822 (VRN1), WO 00/44918(VRN2), WO 99/49064 (GI), WO 00/46358 (FRI), WO 97/29123, WO 99/09174(D8 and Rht), WO 2004/076638, and WO 004/031349 (transcription factors).

Tissue Culture

Further reproduction of the variety can occur by tissue culture andregeneration. Tissue culture of various tissues of celery andregeneration of plants there from is well known and widely published.For example, reference may be had to Teng et al., HortScience. 1992, 27:9, 1030-1032 Teng et al., HortScience. 1993, 28: 6, 669-1671, Zhang etal., Journal of Genetics and Breeding. 1992, 46: 3, 287-290, Webb etal., Plant Cell Tissue and Organ Culture. 1994, 38: 1, 77-79, Curtis etal., Journal of Experimental Botany. 1994, 45: 279, 1441-1449, Nagata etal., Journal for the American Society for Horticultural Science. 2000,125: 6, 669-672, and Ibrahim et al., Plant Cell, Tissue and OrganCulture. (1992), 28(2): 139-145. It is clear from the literature thatthe state of the art is such that these methods of obtaining plants areroutinely used and have a very high rate of success. Thus, anotheraspect of this invention is to provide cells which upon growth anddifferentiation produce celery plants having the physiological andmorphological characteristics of variety TBG 40.

As used herein, the term “tissue culture” indicates a compositioncomprising isolated cells of the same or a different type or acollection of such cells organized into parts of a plant. Exemplarytypes of tissue cultures are protoplasts, calli, plant clumps, and plantcells that can generate tissue culture that are intact in plants orparts of plants, such as embryos, hypocotyls, pollen, flowers, seeds,leaves, stems, roots, root tips, pistils, anthers, meristematic cellsand the like. Means for preparing and maintaining plant tissue cultureare well known in the art. By way of example, a tissue culturecomprising organs has been used to produce regenerated plants. U.S. Pat.Nos. 5,959,185; 5,973,234 and 5,977,445 describe certain techniques, thedisclosures of which are incorporated herein by reference.

Industrial Uses of Celery Cultivar TBG 40

Celery may be used in a variety of manners including but not limited to,use in salads, soups, being filled with cheese, soybean, vegetable,peanut butter or dairy type products, served raw, cooked, baked orfrozen, served as sticks, pieces, diced, or dipped like potato chips, orused as straws.

Tables

In the tables that follow, the traits and characteristics of celerycultivar TBG 40 are given compared to other celery cultivars. Colorreferences made in the Tables refer to the Munsell (MUN) Color Chart.

Table 2 shows the results of a trial comparing characteristics of celerycultivar TBG 40 to celery varieties TBG 39, TBG 42, ADS-1, TBG 29, TallUtah 52-70 R Strain, Tall Utah 52-75, Challenger, Sonora, Conquistador,and Command. The trial was transplanted in Oxnard, Calif. on Aug. 9,2011, and evaluated on Nov. 22, 2011 (105 days). The plant population(58,080 plants to the acre) was higher than the commercial norm ofapproximately 45,000 to 47,000 plants to the acre. Table 2, column 1shows the characteristic and columns 2-12 show the results for TBG 40,TBG 39, TBG 42, ADS-1, TBG 29, Tall Utah 52-70 R Strain, Tall Utah52-75, Challenger, Sonora, Conquistador, and Command, respectively. InTable 2, for the trimmed plant weight, TBG 40=20 cm, TBG 39=15 cm, TBG42=30.5 cm, and the remainder=40 cm. For the number of outer petioles,TBG 40>20 cm, TBG 39>15 cm, TBG 42>30.5 cm, and the remainder>40 cm. Forthe number of inner petioles, TBG 40<20 cm, TBG 39<15 cm, TBG 42<30.5cm, and the remainder <40 cm.

TABLE 2 Tall Utah 52-70 R TBG 40 TBG 39 TBG 42 ADS-1 TBG 29 Strain PlantHeight (cm) Average 42.3 33.4 53.7 82.5 85.2 55.6 Range (40-44) (31-37)(51-57) (76-88) (81-89) (50-63) Whole Plant Average 0.62 0.43 0.98 1.281.54 0.60 Weight (kg) Range (0.43-0.7)  (0.32-0.50) (0.74-1.22)(1.01-1.63) (1.29-1.87) (0.26-0.91) Trimmed Plant Average 0.51 0.30 0.831.00 1.18 0.54 Weight (kg) Range (0.37-0.60) (0.24-0.37) (0.66-1.0) (0.81-1.25) (1.01-1.37) (0.25-0.77) Number of Outer Average 12.6 11.712.5 12.0 12.8 11.7 Petioles Range (10-14) (10-13) (10-14) (10-15)(12-14)  (6-14) Number of Inner Average 7.2 3.2 6.8 7.2 5.1 6.3 PetiolesRange (6-8) (3-4) (5-8) (7-8) (3-7)  (4-10) Length of Outer Average 12.98.4 19.5 31.6 29.1 21.2 Petioles @ joint Range (12-14) (8-9) (18.3-21)  (27.7-34)   (27.7-30.3) (18.7-25.3) (cm) Width of Outer Average 23.425.4 24.3 25.4 28.3 16.8 Petioles @ midrib Range (19.3-28)   (22-28)(21.7-30)   (23.3-28)     (26-31.3) (12.3-21.3) (mm) Thickness of OuterAverage 8.1 8.6 9.8 10.0 0.0 7.7 Petioles @ midrib Range (7-9)   (7-9.7) (8.7-11.3) (9.3-11)  (0-0) (6.7-8.7) (mm) Petiole Color 5gy 5/8 5gy 7/6& 5gy 5/6 5gy 6/6 5gy 6/6 5gy 5/6 (Munsell Color) 5rp 4/8 Leaf Color 5gy4/4 5gy 3/4 5gy 3/4 5gy 3/4 5gy 4/4 5gy 4/6 (Munsell Color) PetioleSmoothness smooth smooth/ smooth smooth smooth rib slight rib PetioleCup cup cup cup cup cup slight cup Tall Utah 52-75 Challenger SonoraConquistador Command Plant Height (cm) Average 69.2 78.1 69.4 74.0 81.5Range (60-75) (62-90) (66-75) (70-81) (76-88) Whole Plant Average 0.811.20 1.02 1.01 1.28 Weight (kg) Range (0.48-1.08) (0.69-1.99)(0.75-1.39) (0.58-1.26) (0.79-1.91) Trimmed Plant Average 0.66 0.92 0.820.80 0.99 Weight (kg) Range (0.42-0.83)  (0.6-1.46) (0.615-1.09) (0.495-1.0)   (0.63-1.43) Number of Outer Average 11.8 10.3 12.3 12.912.2 Petioles Range  (9-14)  (9-12) (11-13) (11-15) (10-15) Number ofInner Average 5.7 5.2 5.6 5.4 5.2 Petioles Range (4-7) (4-7) (4-7) (3-8)(4-6) Length of Outer Average 27.2 32.6 30.2 31.3 31.4 Petioles @ jointRange (23.7-32.3) (25-36) (27.3-32.3) (28.3-34.7) (27.7-37)   (cm) Widthof Outer Average 20.1 23.9 22.3 21.5 23.4 Petioles @ midrib Range(17-24) (20.3-28.7) (20.3-25)   (16.3-27)   (20.7-25.7) (mm) Thicknessof Outer Average 8.1 10.4 8.6 8.6 9.7 Petioles @ midrib Range (6.3-9.7)(8.3-13)  (7.7-9.7) (6.3-10)  (7.3-11)  (mm) Petiole Color 5gy 6/6 5gy5/6 5gy 6/6 5gy 5/6 5gy 7/6 (Munsell Color) Leaf Color 5gy 4/6 5gy 3/45gy 3/4 5gy 4/6 5gy 4/4 (Munsell Color) Petiole Smoothness smooth smoothsmooth smooth smooth Petiole Cup cup cup cup cup cup

As shown in Table 2, celery cultivar TBG 40 was taller than TBG 39, butshorter than the rest of the other celery varieties in the trial. TBG 40also had a high number of outer and inner petioles.

Table 3 shows the results of a trial comparing characteristics of celerycultivar TBG 40 to celery varieties TBG 39, TBG 41, TBG 42, ADS-1, TBG29, ADS-20, Challenger, Sonora, Conquistador, Command, and Mission. Thetrial was transplanted in Oxnard, Calif. on Mar. 14, 2013, and evaluatedon Jun. 21, 2013 (99 days). The plant population (58,080 plants to theacre) was higher than the commercial norm of approximately 45,000 to47,000 plants to the acre. Table 3, column 1 shows the characteristicand columns 2-13 show the results for TBG 40, TBG 39, TBG 41, TBG 42,ADS-1, TBG 29, ADS-20, Challenger, Sonora, Conquistador, Command, andMission, respectively. In Table 3, for the trimmed plant weight, TBG40=20 cm, TBG 39=10 cm, TBG 41=18 cm, TBG 42=25 cm, and the remainder=40cm. For the number of outer petioles, TBG 40>20 cm, TBG 39>10 cm, TBG41>18 cm, TBG 42>25 cm, and the remainder>40 cm. For the number of innerpetioles, TBG 40<20 cm, TBG 39<10 cm, TBG 41<18 cm, TBG 42<25 cm, andthe remainder <40 cm.

TABLE 3 TBG 40 TBG 39 TBG 41 TBG 42 ADS-1 TBG 29 Plant Height Average42.2 31.7 39.8 55.8 72.4 75.4 (cm) Range (39-46) (28-35) (36-43) (54-60)(69-76) (71-81) Whole Plant Average 0.9 0.5 0.6 1.3 1.4 1.5 Weight (kg)Range  (0.7-0.96) (0.44-0.66) (0.44-0.71) (1.08-1.47) (1.28-1.68)(1.32-1.67) Trimmed Plant Average 0.72 0.31 0.44 1.03 1.18 1.24 Weight(kg) Range (0.58-0.81) (0.24-0.37) (0.35-0.54) (0.83-1.2)  (1.01-1.37)(1.08-1.38) Number of Average 14.2 13.6 13.1 13.6 12.8 14.7 OuterPetioles Range (12-16) (11-17) (10-15) (12-15) (12-14) (13-16) Number ofAverage 7.6 2.8 4.0 7.3 7.7 6.9 Inner Petioles Range (6-9) (2-5) (3-5)(6-9)  (6-10) (5-8) Length of Average 13.2 7.1 12.8 20.7 27.8 27.3 OuterPetioles Range (12.3-14)     (6-8.3) (11.7-13.7) (19.3-21.7) (24.3-30.7)(24.7-29)   joint (cm) Width of Outer Average 36.3 49.5 26.1 28.5 30.932.8 Petioles @ Range (33-38)  (30.7-181.3)   (23-29.3) (26.3-33.3)(27.7-34)   (30-36) midrib (mm) Thickness of Average 8.5 9.0 9.4 11.510.9 11.1 Outer Petioles Range (8-9) (7.7-9.7)   (7-11.3) (10.3-12)  (9.7-12)  (10-12) @ midrib (mm) Petiole Color 5gy 6/6 2.5r 5/6- 5gy 5/65gy 5/6 5gy 6/6 5gy 5/6 (Munsell Color ) 5gy 6/6 Leaf Color 5gy 3/4 5gy3/4 5gy 3/4 5gy 3/4 5gy 3/4 5gy 3/4 (Munsell Color) Petiole smooth/slight rib smooth/ smooth smooth smooth/ Smoothness slight rib slightrib slight rib Petiole Cup slight cup cup slight cup/ cup cup slight cupcup ADS-20 Challenger Sonora Conquistador Command Mission Plant HeightAverage 72.8 83.8 74.9 74.9 79.4 79.1 (cm) Range (69-77) (76-95) (68-79)(68-80) (72-83) (74-88) Whole Plant Average 1.1 1.4 1.0 1.2 1.5 1.4Weight (kg) Range (0.82-1.4)  (1.19-1.72) (0.74-1.31) (0.76-1.57)(1.24-1.82) (0.92-1.71) Trimmed Plant Average 0.87 1.13 0.83 0.98 1.241.11 Weight (kg) Range (0.66-1.1)  (0.93-1.38)  (0.6-1.06)  (0.6-1.23)(0.98-1.52) (0.66-1.31) Number of Average 10.1 12.4 12.6 12.8 12.3 13.7Outer Petioles Range  (9-13) (11-14) (11-14) (10-15) (10-14) (13-15)Number of Average 4.9 6.1 5.7 5.8 7.1 6.9 Inner Petioles Range (3-6)(5-8) (5-7) (3-7) (5-8) (5-9) Length of Average 32.0 36.3 31.4 31.4 28.433.0 Outer Petioles Range (28.7-35.3) (30.3-42)   (29.3-33)   (27-34)(24.3-33.3)   (29-38.3) joint (cm) Width of Outer Average 26.4 26.2 24.725.2 27.2 28.0 Petioles @ Range   (23-28.7) (24.7-28.7) (21.3-26.7)(20.3-28.7)   (24-32.7) (21.7-32.3) midrib (mm) Thickness of Average 9.810.6 8.9 9.3 10.3 9.6 Outer Petioles Range  (8.7-10.7)  (9.7-11.7)  (8-9.7) (8.7-10)   (9.7-11.3)  (7-11) @ midrib (mm) Petiole Color 5gy6/6 5gy 6/6 5gy 6/6 5gy 6/6 5gy 7/6 5gy 7/6 (Munsell Color ) Leaf Color5gy 3/4 5gy 3/4 5gy 3/4 5gy 3/4 5gy 4/4 5gy 4/4 (Munsell Color) Petiolesmooth/ smooth/ smooth/ smooth/ smooth slight rib Smoothness slight ribslight rib slight rib slight rib Petiole Cup cup cup slight cup/ cup cupcup cup

As shown in Table 3, celery cultivar TBG 40 was taller than TBG 39 andTBG 41, but was shorter than the rest of the varieties in the trial. TBG40 also had a high number of inner and outer petioles compared to theother varieties and wider petioles than all other varieties except TBG39.

Table 4 shows the results of a trial comparing characteristics of celerycultivar TBG 40 to celery varieties TBG 39, TBG 41, ADS-1, TBG 29,ADS-20, Tall Utah 5270 R Strain, Tall Utah 5275, Challenger, Sonora,Conquistador, Command, and Mission. The trial was transplanted inOxnard, Calif. on Mar. 14, 2017, and evaluated on Jun. 22, 2017 (100days). The plant population (58,080 plants to the acre) was higher thanthe commercial norm of approximately 45,000 to 47,000 plants to theacre. Table 4, column 1 shows the characteristic and columns 2-14 showthe results for TBG 40, TBG 39, TBG 41, ADS-1, TBG 29, ADS-20, Tall Utah5270 R Strain, Tall Utah 5275, Challenger, Sonora, Conquistador,Command, and Mission, respectively.

TABLE 4 Tall Utah 5270 R TBG 40 TBG 39 TBG 41 ADS-1 TBG 29 ADS-20 StrainPlant Height Avg. 40.0 30.0 36.6 81.5 80.6 69.4 73.9 (cm) Range (38-42)(28-32) (34-40) (79-84) (76-87) (20-80) (71-77) Whole Plant Avg. 0.610.50 0.49 1.59 1.90 1.24 1.14 Weight (kg) Range (0.49-0.80) (0.44-0.58)(0.36-0.58) (1.46-1.79) (1.54-2.26)   (1-1.44) (0.78-1.68) Trimmed PlantAvg. 0.59 0.50 0.49 1.26 17.72 1.00 0.91 Weight (kg) Range (0.49-0.68)(0.44-0.58) (0.36-0.58) (1.13-1.47) (1.23-1.64) (0.78-1.14) (0.61-1.35)(all stalks cut at 40 cm) Number of Avg. 1.9 0.0 0.0 13.3 15.8 11.8 11.7Outer Petioles Range (0-4) (0-0) (0-0) (12-15) (13-18) (10-13) (10-15)(>40 cm) Number of Avg. 16.5 15.5 17.9 8.2 6.6 5.4 7.1 Inner PetiolesRange (14-20) (13-17) (14-21) (7-9) (5-8) (5-6)  (6-10) (<40 cm) Lengthof Outer Avg. 12.4 8.4 11.7 33.1 29.9 31.5 21.7 Petioles @ joint Range(11.3-14.3)   (8-9.3)   (10-14.3) (31-35) (26.7-33.7) (28.3-33)     (0-32.7) (cm) Width of Outer Avg. 28.6 28.1 24.1 29.4 33.0 25.8 17.2Petioles @ Range (25.7-33.3) (20.3-31)   (21.7-26)   (26.7-33.3)(29.7-37.3) (22-30)   (0-30.3) midrib (mm) Thickness of Avg. 9.3 9.8 9.811.2 11.5 10.3 6.7 Outer Petioles Range    (8-10.3)   (9-10.7) (8.7-10.3) (10-12) (10.7-12.3) (9.7-12)    (0-11.3) @ midrib (mm)Petiole Color 5gy 7/6 5gy 5/4- 5gy 6/6 5gy 5/6 5gy 5/6- 5gy 6/6 5gy 7/6(Munsell Color) 5gy 7/6- 5gy 6/6 5rp 6/8 Leaf Color 5gy 3/4 5gy 3/4 5gy3/4 5gy 3/4 5gy 3/4 5gy 3/4 5gy 3/4 (Munsell Color) Petiole smoothslight rib/ smooth smooth/ smooth/ slight rib rib Smoothness rib slightrib slight rib Petiole Cup slight cup slight slight cup slight cupslight cup/cup cup/cup cup/cup cup Tall Utah Chall- Conqui- 5275 engerSonora stador Command Mission Plant Height Avg. 74.1 81.5 75.4 77.5 77.378.5 (cm) Range (69-78) (72-89) (72-82) (68-83) (71-85) (74-83) WholePlant Avg. 1.30 1.41 1.48 1.36 1.36 1.63 Weight (kg) Range (0.98-1.65)(1.18-1.63) (1.17-2.05) (0.99-1.9)  (0.87-1.68) (1.08-2)   Trimmed PlantAvg. 1.02 1.08 1.13 1.04 1.09 1.22 Weight (kg) Range (0.77-1.34)(0.93-1.2)  (0.88-1.57) (0.73-1.48)  (0.7-1.33) (0.84-1.47) (all stalkscut at 40 cm) Number of Avg. 12.2 11.4 13.0 15.5 14.0 15.1 OuterPetioles Range (10-14)  (9-13) (12-16) (13-18) (10-17) (12-17) (>40 cm)Number of Avg. 7.0 6.3 7.1 6.8 6.9 7.0 Inner Petioles Range (5-9) (5-8) (5-10) (5-8) (6-8) (5-9) (<40 cm) Length of Outer Avg. 30.1 31.6 30.832.7 30.9 33.2 Petioles @ joint Range   (26-33.3) (26-35) (28-34)(30.7-34)   (28.7-32.7) (30.3-37.7) (cm) Width of Outer Avg. 26.2 25.328.3 24.5 23.9 24.5 Petioles @ Range (23.3-28.3)  (0-31) (23.3-34)    (19-31.7) (22-27) (15.7-28)   midrib (mm) Thickness of Avg. 10.7 10.69.4 8.9 10.3 10.1 Outer Petioles Range   (9-12.3)  (0-14) (8.3-10) (7.7-10)   (9.7-11.3)  (8.7-11.3) @ midrib (mm) Petiole Color 5gy 7/6-5gy 7/4- 5gy 6/6 7gy 7/6 5gy 7/6 5gy 6/6 (Munsell Color) 5gy 6/6 5gy 7/6Leaf Color 5gy 3/4 5gy 4/4 5gy 3/4 5gy 3/4 5gy 3/4 5gy 3/4 (MunsellColor) Petiole smooth smooth/ smooth/ rib smooth/ smooth Smoothnessslight rib slight rib slight rib Petiole Cup slight cup slight cup cupcup cup/cup cup/cup

As shown in Table 4, celery cultivar TBG 40 was taller than TBG 39 andTBG 41, but was shorter than the rest of the varieties in the trial. TBG40 also the second highest number of inner petioles and the third widestpetioles compared to the other varieties.

Tables 5A and 5B show the results of trials comparing the boltingcharacteristics of celery cultivar TBG 40 to celery varieties TBG 39,TBG 41, TBG 42, ADS-1, TBG 29, ADS-20, Tall Utah 52-70 ‘It’ Strain, TallUtah 52-75, Challenger, Sonora, Conquistador, Command, and Mission. Thedifferent trials were grown in the prominent bolting windows in 2008,2009, 2010, 2011, 2014, 2015, 2016, 2017, and 2018. Santa Paula wasselected for production of more recent trials because it is not in theprominent coastal plain of Ventura County, California where most WestCoast celery is grown this time of year in order to minimize coldaccumulation and the initiation of bolting (development of seed stems).Santa Paula is inland where is there is less warming due to its distancefrom the warming influence of the Pacific Ocean. Measurements were forthe length of the seed stem developed in the celery plant and morebolting tolerant are considered those varieties with the least amount ofseed stem development. In Tables 5A and 5B, the trial header shows thelocation for each trial, the harvest date, and the number of hours below50° F. to which the celery was exposed. Seed stem length is shown incentimeters. Blanks in the data (NA) occur where the particular cultivarwas not included in the trial and data is ‘not available’.

TABLE 5A Santa Paula, CA Santa Paula, CA Santa Paula, CA Harvest: May 7,2018 Harvest: May 3, 2017 Harvest: Apr. 25, 2016 (140 days) (128 days)(129 days) 455 hours below 50° F. 608 hours below 50° F. 891 hours below50° F. Average Range Median Average Range Median Average Range TBG 40 NANA NA NA NA NA NA NA TBG 39 16.3 (12-23) 16.0 NA NA NA NA NA TBG 41 0.6(0-2) 0.5 0.0 (0-0) 0.0 0.1   (0-0.5) TBG 42 NA NA NA 3.8  (0-14) 2.017.0 (13-20) ADS-1 24.3  (5-43) 28.5 9.9  (1-23) 7.8 16.4  (9-26) TBG 2930.7  (8-57) 29.5 10.5 (1.5-22)  9.8 26.0 (16-39) ADS-20 0.4   (0-0.5)0.5 0.0 (0-0) 0.0 0.0 (0-0) Tall Utah 52-70 70.9 (53-87) 70.5 49.8(34-67) 48.0 NA NA ‘R’ Strain Tall Utah 52-75 44.2 (10-65) 47.5 28.5(9.5-54)  34.0 40.8 (25-57) Challenger 40.3 (23-55) 39.5 20.3(13.5-25.5) 21.1 30.2 (22-38) Sonora 16.2 (0.5-32)  14.0 6.7 (1.5-17) 5.3 29.0  (7-41) Conquistador 28.4  (0-52) 31.5 15.6 (7.5-40)   18.030.1 (19-40) Command 27.1  (5-57) 24.5 10.2  (1.5-24.5) 9.8 29.7 (19-40)Mission 26.6  (6-68) 19.0 13.6    (2-28.2) 13.3 22.1 (12-32) SantaPaula, CA Santa Paula, CA Santa Paula, CA Harvest: Apr. 25, 2016Harvest: Apr. 28, 2015 Harvest: Apr. 14, 2014 (129 days) (125 days) (125days) 891 hours below 50° F. 848 hours below 50° F. 895 hours below 50°F. Median Average Range Median Average Range Median TBG 40 NA NA NA NANA NA NA TBG 39 NA NA NA NA NA NA NA TBG 41 0.0 0.0 (0-0) 0.0 NA NA NATBG 42 17.0 2.0 (0-7) 1.1 5.4 (2-9) 5.5 ADS-1 16.5 8.1 (0.2-19)  8.0 4.1(0-9) 4.5 TBG 29 25.5 3.5 (0.2-10)  2.0 2.8 (0-7) 2.0 ADS-20 0.0 0.1  (0-0.3) 0.0 0.0 (0-0) 0.0 Tall Utah 52-70 NA NA NA NA 19.5 (13-28)18.0 ‘R’ Strain Tall Utah 52-75 41.0 NA NA NA 18.1 (10-33) 14.5Challenger 31.0 NA NA NA 12.7 (10-15) 12.5 Sonora 32.0 10.3  (1-20) 10.512.7  (5-18) 12.0 Conquistador 29.5 7.2  (0-19) 5.0 10.1  (4-17) 11.0Command 29.5 18.9  (5-32) 15.5 11.1  (7-18) 10.5 Mission 23.0 3.4 (0-15) 1.3 5.1 (0.5-9)   5.0

TABLE 5B Santa Paula, CA Oxnard, CA Santa Paula, CA Oxnard, CA Harvest:Apr. 22, 2011 Harvest: May 15, 2010 Harvest: Apr. 22, 2009 Harvest: May1, 2008 1094 hours below 50° F. 835 hours below 50° F. 908 hours below50° F. 901 hours below 50° F. Average Range Median Average Range MedianAverage Range Median Average Range Median TBG 40 NA NA NA NA NA NA 23.3 (0-37) 27.5 NA NA NA TBG 39 11.8  (6-18) 11.0 NA NA NA NA NA NA 14.8(12-18) 15.5 TBG 41 NA NA NA 0.5 (0-2) 0.0 9.6  (0-26) 7.5 NA NA NA TBG42 13.3  (8-19) 13.5 NA NA NA NA NA NA 2.2 (0-7) 0.3 ADS-1 20.2  (8-27)21.0 13.1  2-35 12.0 45.3 21-67 46.5 24.5 13-38 25.0 TBG 29 NA NA NA NANA NA NA NA NA NA NA NA ADS-20 0.7 0-2 0.5 0 0-0 0.0 0 0-0 0.0 0   0-0.50.0 Tall Utah 52-70 38.4 (28-53) 39.5 39.2 26-55 37.5 75.3 70-82 75.548.9 39-58 49.0 ‘R’ Strain Tall Utah 52-75 NA NA NA 19  1-42 18 58.844-73 58.5 37.9 33-47 37 Challenger 25.5 (16-35) 25.0 NA NA NA NA NA NA36.0 30-48 33.5 Sonora 19.2 (12-32) 18.0 12.5  3-27 11.0 50.3 36-56 55.022.4 17-31 22.5 Conquistador 13.4 (11-21) 13.0 NA NA NA 60.8 50-83 59.029.3 22-39 28.5 Command 21.7  (8-31) 22.0 NA NA NA NA NA NA 27.1 20-3725.5 Mission NA NA NA NA NA NA NA NA NA 31.7 12-53 32

As shown in Table 5B, in the 2009 bolting trial, celery cultivar TBG 40had a shorter seed stem length than all other varieties except for TBG41.

The use of the terms “a,” “an,” and “the,” and similar referents in thecontext of describing the invention (especially in the context of thefollowing claims) are to be construed to cover both the singular and theplural, unless otherwise indicated herein or clearly contradicted bycontext. The terms “comprising,” “having,” “including,” and “containing”are to be construed as open-ended terms (i.e., meaning “including, butnot limited to,”) unless otherwise noted. Recitation of ranges of valuesherein are merely intended to serve as a shorthand method of referringindividually to each separate value falling within the range, unlessotherwise indicated herein, and each separate value is incorporated intothe specification as if it were individually recited herein. Forexample, if the range 10-15 is disclosed, then 11, 12, 13, and 14 arealso disclosed. All methods described herein can be performed in anysuitable order unless otherwise indicated herein or otherwise clearlycontradicted by context. The use of any and all examples, or exemplarylanguage (e.g., “such as”) provided herein, is intended merely to betterilluminate the invention and does not pose a limitation on the scope ofthe invention unless otherwise claimed. No language in the specificationshould be construed as indicating any non-claimed element as essentialto the practice of the invention.

DEPOSIT INFORMATION

A deposit of the A. Duda & Sons, Inc. proprietary celery cultivar TBG 40disclosed above and recited in the appended claims has been made withthe American Type Culture Collection (ATCC), 10801 University Boulevard,Manassas, Virginia 20110 under the terms of the Budapest Treaty. Thedate of deposit was December 2, 2020. The deposit of 625 seeds was takenfrom the same deposit maintained by A. Duda & Sons, Inc. since prior tothe filing date of this application. All restrictions will beirrevocably removed upon granting of a patent, and the deposit isintended to meet all of the requirements of 37 C.F.R. §§ 1.801-1.809.The ATCC Accession Number is PTA-126888. The deposit will be maintainedin the depository for a period of thirty years, or five years after thelast request, or for the enforceable life of the patent, whichever islonger, and will be replaced as necessary during that period.

While a number of exemplary aspects and embodiments have been discussedabove, those of skill in the art will recognize certain modifications,permutations, additions, and sub-combinations thereof. It is thereforeintended that the following appended claims and claims hereafterintroduced are interpreted to include all such modifications,permutations, additions, and sub-combinations as are within their truespirit and scope.

What is claimed is:
 1. A seed, plant, or a plant part thereof, of celerycultivar designated TBG 40, wherein a representative sample of seed ofsaid cultivar has been deposited under ATCC Accession No. PTA-126888. 2.A celery plant having all of the physiological and morphologicalcharacteristics of the celery plant of claim 1, or a plant part thereof.3. A tissue or cell culture produced from protoplasts or cells from theplant of claim 1, wherein said cells or protoplasts are produced from aplant part selected from the group consisting of leaf, callus, pollen,ovule, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip,pistil, anther, flower, seed, shoot, stem, stalk, petiole and sucker. 4.A celery plant regenerated from the tissue culture of claim 3, whereinsaid regenerated plant comprises all of the morphological andphysiological characteristics of celery cultivar TBG
 40. 5. A method ofproducing a celery seed, wherein the method comprises crossing the plantof claim 1 with a different celery plant and harvesting the resultantcelery seed.
 6. An F₁ celery seed produced by the method of claim
 5. 7.An F₁ celery plant, or a plant part thereof, produced by growing saidseed of claim
 6. 8. A method of producing an herbicide resistant celeryplant, wherein said method comprises introducing a gene conferringherbicide resistance into the plant of claim
 1. 9. An herbicideresistant celery plant produced by the method of claim 8, wherein thegene confers resistance to a herbicide selected from the groupconsisting of glyphosate, sulfonylurea, imidazolinone, dicamba,glufosinate, phenoxy proprionic acid, L-phosphinothricin, cyclohexone,cyclohexanedione, triazine, benzonitrile protoporphyrinogen oxidase(PPO)-inhibitor herbicides, auxin herbicides, and broxynil, wherein saidplant comprises said gene and otherwise comprises all of thephysiological and morphological characteristics of celery cultivar TBG40.
 10. A method of producing a pest or insect resistant celery plant,wherein said method comprises introducing a gene conferring pest orinsect resistance into the celery plant of claim
 1. 11. A pest or insectresistant celery plant produced by the method of claim 10, wherein saidplant comprises said gene and otherwise comprises all of thephysiological and morphological characteristics of celery cultivar TBG40.
 12. The celery plant of claim 11, wherein the gene encodes aBacillus thuringiensis (Bt) endotoxin, wherein said plant comprises saidgene and otherwise comprises all of the physiological and morphologicalcharacteristics of celery cultivar TBG
 40. 13. A method of producing adisease resistant celery plant, wherein said method comprisesintroducing a gene into the celery plant of claim
 1. 14. A diseaseresistant celery plant produced by the method of claim 13, wherein saidplant comprises said gene and otherwise comprises all of thephysiological and morphological characteristics of celery cultivar TBG40.
 15. A method of producing a celery plant with modified fatty acidmetabolism or modified carbohydrate metabolism comprising transformingthe celery plant of claim 1 with a transgene encoding a protein selectedfrom the group consisting of fructosyltransferase, levansucrase,a-amylase, invertase and starch branching enzyme or DNA encoding anantisense of stearyl-ACP desaturase.
 16. A celery plant having modifiedfatty acid metabolism or modified carbohydrate metabolism produced bythe method of claim 15, wherein said plant comprises said transgene andotherwise comprises all of the physiological and morphologicalcharacteristics of celery cultivar TBG
 40. 17. A method for producing amale sterile celery plant, wherein said method comprises transformingthe celery plant of claim 1 with a nucleic acid molecule that confersmale sterility.
 18. A male sterile celery plant produced by the methodof claim 17, wherein said plant comprises said nucleic acid andotherwise comprises all of the physiological and morphologicalcharacteristics of celery cultivar TBG
 40. 19. A method of introducing adesired trait into celery cultivar TBG 40, wherein the method comprises:(a) crossing a TBG 40 plant, wherein a representative sample of seed wasdeposited under ATCC Accession No. PTA-126888, with a plant of anothercelery cultivar that comprises a desired trait to produce progenyplants, wherein the desired trait is selected from the group consistingof improved nutritional quality, industrial usage, male sterility,herbicide resistance, insect resistance, modified seed yield, modifiedlodging resistance, modified iron-deficiency chlorosis, resistance tobacterial disease, resistance to fungal disease, and resistance to viraldisease; (b) selecting one or more progeny plants that have the desiredtrait to produce selected progeny plants; (c) backcrossing the selectedprogeny plants with a TBG 40 plant to produce backcross progeny plants;(d) selecting for backcross progeny plants that have the desired trait;and (e) repeating steps (c) and (d) two or more times in succession toproduce selected third or higher backcross progeny plants that comprisethe desired trait.
 20. A celery plant produced by the method of claim19, wherein the plant has the desired trait and otherwise all of thephysiological and morphological characteristics of celery cultivar TBG40.
 21. The celery plant of claim 20, wherein the desired trait isherbicide resistance and the resistance is conferred to a herbicideselected from the group consisting of imidazolinone, dicamba,cyclohexanedione, sulfonylurea, glyphosate, glufosinate, phenoxyproprionic acid, L-phosphinothricin, triazine, benzonitrileprotoporphyrinogen oxidase (PPO)-inhibitor herbicides, auxin herbicides,and broxynil.
 22. The celery plant of claim 20, wherein the desiredtrait is insect resistance and the insect resistance is conferred by agene encoding a Bacillus thuringiensis endotoxin.
 23. The celery plantof claim 20, wherein the desired trait is male sterility and the traitis conferred by a cytoplasmic nucleic acid molecule.
 24. A method ofproducing a genetically modified celery plant, wherein the methodcomprises mutation, transformation, gene conversion, genome editing, RNAinterference or gene silencing of the plant of claim
 1. 25. Agenetically modified celery plant produced by the method of claim 24,wherein the plant comprises the genetic modification and otherwisecomprises all of the physiological and morphological characteristics ofcelery cultivar TBG 40.